Compounds

ABSTRACT

This disclosure relates to compounds of formula (I), which are modulators of STING. Also disclosed are pharmaceutical compositions comprising compounds of formula (I) and methods of using compounds of formula (I) in the treatment or prevention of diseases ameliorated by the modulation of STING.

This application claims priority to United Kingdom provisional application no. GB1918761.6 (filed 18 Dec. 2019) and United Kingdom provisional application no. GB2009944.6 (filed 30 Jun. 2020). The entire contents of each of GB1918761.6 and GB2009944.6 are incorporated herein by reference.

The present invention relates to substituted benzimidadzole dimers and their use as pharmaceuticals, and in particular, in treating diseases ameliorated by the modulation of STING.

BACKGROUND TO THE INVENTION

Vertebrates are constantly threatened by the invasion of microorganisms and have evolved mechanisms of immune defense to eliminate infective pathogens. In mammals, this immune system comprises two branches; innate immunity and adaptive immunity. The innate immune system is the first line of defense which is initiated by Pattern Recognition Receptors (PRRs) which detect ligands from the pathogens as well as damage associated molecular patterns (Takeuchi 2010). A growing number of these receptors have been identified including Toll-like receptors (TLRs), C-type lectin receptors, retinoic acid inducible gene I (RIG-I)-like receptors and NOD-like receptors (NLRs) and also double stranded DNA sensors. Activation of PRRs leads to up-regulation of genes involved in the inflammatory response including type 1 interferons, pro-inflammatory cytokines and chemokines which suppress pathogen replication and facilitate adaptive immunity.

The adaptor protein STING (Stimulator of Interferon Genes), also known as TMEM 173, MPYS, MITA and ERIS, has been identified as a central signalling molecule in the innate immune response to cytosolic nucleic acids (Ishikawa 2008; WO2013/166000). Activation of STING results in up-regulation of IRF3 and NFκB pathways leading to induction of type 1 interferons including Interferon-β and other cytokines. STING is critical for responses to cytosolic DNA of pathogen or host origin, and of unusual nucleic acids called Cyclic Dinucleotides (CDNs).

CDNs were first identified as bacterial secondary messengers responsible for controlling numerous responses in the prokaryotic cell. Bacterial CDNs, such as c-di-GMP, are symmetrical molecules characterized by two 3′,5′ phosphodiester linkages.

Direct activation of STING by bacterial CDNs has recently been confirmed through X-ray crystallography (Burdette 2013). Bacterial CDNs and their analogues have consequently attracted interest as potential vaccine adjuvants (Libanova 2012; WO2007/054279; WO2005/087238).

More recently, the response to cytosolic DNA has been elucidated and shown to involve generation, by an enzyme called cyclic GMP-AMP synthase (cGAS, previously known as C₆orfl50 or MB21 D1), of a novel mammalian CDN signalling molecule identified as cGAMP, which then activates STING. Unlike bacterial CDNs, cGAMP is an asymmetrical molecule characterized by its mixed 2′,5′ and 3′,5′ phosphodiester linkages (Gao 2013A). Interaction of cGAMP (II) with STING has also been demonstrated by X-ray crystallography (Cai 2014).

Interferon was first described as a substance which could protect cells from viral infection (Isaacs 1957). In man, the type I interferons are a family of related proteins encoded by genes on chromosome 9 and encoding at least 13 isoforms of interferon alpha (IFNα) and one isoform of interferon beta (IFNβ). Recombinant IFNα was the first approved biological therapeutic and has become an important therapy in viral infections and in cancer. As well as direct antiviral activity on cells, interferons are known to be potent modulators of the immune response, acting on cells of the immune system.

Administration of a small molecule compound which could modulate the innate immune response, including the activation or inhibition of type I interferon production and other cytokines, could become an important strategy for the treatment or prevention of human diseases including viral infections and autoimmune disease. This type of immunomodulatory strategy has the potential to identify compounds which may be useful not only in infectious diseases but also in cancer (Zitvogel 2015), allergic diseases (Moisan 2006), neurodegenerative diseases such as amyotrophic lateral sclerosis and multiple sclerosis (Lemos 2014; Cirulli 2015; Freischmidt 2015), other inflammatory conditions such as irritable bowel disease (Rakoff-Nahoum 2004), and as vaccine adjuvants (Persing 2002; Dubensky 2013).

STING is essential for antimicrobial host defense, including protection against a range of DNA and RNA viruses and bacteria (reviewed in McNab 2015; Ma 2016). Herpesviridae, Flaviviridae, Coronaviridae, Papillomaviridae, Adenoviridae, Hepadnaviridae, ortho- and paramyxoviridae and rhabdoviridae have evolved mechanisms to inhibit STING mediated Type I interferon production and evade host immune control (Holm 2016; Ma 2015; Wu 2015; Liu 2016; Chen 2014; Lau 2013; Ding 2013; Nitta 2013; Sun 2012; Aguirre 2012; Ishikawa 2009). Thus, small molecule activation of STING could be beneficial for treatment of these infectious diseases.

In contrast, increased and prolonged type I IFN production is associated with a variety of chronic infections, including Mycobacteria (Collins 2015; Wassermann 2015; Watson 2015), Franciscella (Storek 2015; Jin 2011A), Chlamydia (Prantner 2010), Plasmodium (Sharma 2011), and HIV (Herzner 2015; Gao 2013B). Similarly, excess type I interferon production is found among patients with complex forms of autoimmune disease. Genetic evidence in humans and support from studies in animal models support the hypothesis that inhibition of STING results in reduced type I interferon that drives autoimmune disease (Crow 2006; Stetson 2008). Therefore, inhibitors of STING provide a treatment to patients with chronic type I interferon and proinflammatory cytokine production associated with infections or complex autoimmune diseases. Allergic diseases are associated with a Th2-biased immune-response to allergens. Th2 responses are associated with raised levels of IgE, which, via its effects on mast cells, promotes a hypersensitivity to allergens, resulting in the symptoms seen, for example, in allergic rhinitis and asthma. In healthy individuals the immune-response to allergens is more balanced with a mixed Th2/Th1 and regulatory T cell response. Induction of Type 1 interferons have been shown to result in reduction of Th2-type cytokines in the local environment and promote Th1/Treg responses. In this context, induction of type 1 interferons by, for example, activation of STING, may offer benefit in treatment of allergic diseases such as asthma and allergic rhinitis (Huber 2010).

Compounds that bind to STING and act as agonists have been shown to induce type 1 interferons and other cytokines on incubation with human PBMCs. Compounds which induce human interferons may be useful in the treatment of various disorders, for example the treatment of allergic diseases and other inflammatory conditions for example allergic rhinitis and asthma, the treatment of infectious diseases, neurodegenerative disease, pre-cancerous syndromes and cancer, and may also be useful as immugenic composition or vaccine adjuvants. Compounds that bind to STING may act as antagonists and could be useful in the treatment, for example of autoimmune diseases. It is envisaged that targeting STING with activation or inhibiting agents may be a promising approach for preventing and treating diseases and conditions in which modulation for the type 1 IFN pathway is beneficial, including inflammatory, allergic and autoimmune diseases, infectious diseases, cancer, pre-cancerous syndromes and as immugenic compositions or vaccine adjuvants.

Skin cancers and various skin viral infections involve immune privileged environment and activation of local immune response to the lesions may be a topical therapeutic approach. STING agonists may be used for treating viral warts, superficial skin cancers and premalignant actinic keratoses. By a dual mechanism of action, STING activation (e.g., via microneedle patch delivery or topical formulation) may be used to control HPV directly via antiviral type I interferon production and indirectly by enhancing the adaptive immune response downstream of innate immune activation. STING agonist can activate the innate immune response in the lesion and drive the anti-HPV T-cell response.

Recent evidence has indicated that spontaneous activation of the STING pathway within tumor-resident dendritic cells leads to type I IFN production and adaptive immune responses against tumors. Furthermore, activation of this pathway in antigen presenting cells (APCs) within the tumor microenvironment drives the subsequent T-cell priming against tumor-associated antigens (Corrales 2015). International Patent Applications WO2014/093936, WO2014/189805, WO2013/185052, WO2015/077354 and WO2015/185565 disclose certain cyclic di-nucleotides and their use in inducing an immune response via activation of STING.

Additionally non-CDN compounds have been described as active agonists of STING.

Applications WO2019/069269, WO2019/069270, WO2017/175156, and WO2017/175147 and Ramanjulu 2018 describe certain amidobenzimidazole-based and diamidobenzimidazole-based compounds and their use in modulation of STING.

Applications WO2019/027858 and US2018/0093964 describe certain benzo[b]thiophene compounds and their use as agonists of STING. WO2019/195063 describe Aza-benzothiophene compounds, and WO2019/195124 describes benzothiophenes and related compounds and their use as agonists of STING. WO 2019/219820 describes benzothiophenes, thienopyridines and thienopyrimidines as modulators of STING.

Applications WO2018/234808, WO2018/234807, GB2563642A, WO2018/234805 describe certain arylamido compounds and their use as modulators of STING.

Application WO2019/165032 describes certain imidazole-pyridazine compounds and their use as agonists of STING.

STING has known polymorphisms, including alleles encoding histidine at position 232, which are refractory to bis-3′,5′ (canonical) CDNs, but not 2′,5′-3′,5′ (non-canonical, mixed linkage) CDNs (Diner 2013; Jin 2011B). Single nucleotide polymorphisms in the STING gene have been reported to affect the responsiveness to bacterial-derived canonical CDNs (Diner 2013; Gao 2013C; Conlon 2013). Five major haplotypes of STING have been reported (WT, R232H, HAQ, AQ and Q alleles), which vary at amino acid positions 71, 230, 232 and 293 (Jin 2011B; Yi 2013).

The compounds of this invention modulate the activity of STING, and accordingly, may provide a beneficial therapeutic impact in the prevention and treatment of diseases, disorders and/or conditions in which modulation of STING (Stimulator of Interferon Genes) is beneficial, for example for inflammation, allergic and autoimmune diseases, infectious diseases, cancer, pre-cancerous syndromes and as vaccine adjuvants.

All publications, patents and patent applications that may be cited herein are hereby incorporated by reference in their entirety.

Any discussion of documents, acts, materials, devices, articles or the like which has been included in the present specification is not to be taken as an admission that any or all of these matters form part of the prior art base or were common general knowledge in the field relevant to the present disclosure as it existed before the priority date of each claim of this application.

Any embodiment herein shall be taken to apply mutatis mutandis to any other embodiment unless specifically stated otherwise.

The present disclosure is not to be limited in scope by the specific embodiments described herein, which are intended for the purpose of exemplification only. Functionally-equivalent products, compositions and methods are clearly within the scope of the invention, as described herein.

SUMMARY OF THE INVENTION

A first aspect of the present invention provides a compound of formula I:

wherein: Y is either (CH₂)_(n), where n is from 2 to 4, or —CH₂—CH═CH—CH₂—; R¹ and R¹¹ are independently selected from the group consisting of: —C(═O)OH, —C(═O)OR^(P1), Br, F, tetrazolyl, oxo-oxadiazolyl and

(2H-triazol-4-yl), where R^(P1) is selected from methyl and ethyl; A¹ is CR^(A) or N; A² is CR^(B) or N; A³ is CR^(C) or N; A⁴ is CR^(D) or N; where no more than two of A¹, A², A³, and A⁴ may be N; one, two or three of R^(A), R^(B), R^(C) and R^(D), (if present) are selected from H, F, Cl, Br, I, Me, Et, CF₃, cyclopropyl, cyano, OMe, OEt, CH₂OH, CH₂OMe and OH; the remainder of R^(A), R^(B), R^(C) and R^(D), (if present) are H; A¹¹ is CR^(AA) or N; A¹² is CR^(BB) or N; A¹³ is CR^(CC) or N; A¹⁴ is CR^(DD) or N; where no more than two of A¹¹, A¹², A¹³ and A¹⁴ may be N; one, two or three of R^(AA), R^(BB), R^(CC) and R^(DD), (if present) are selected from H, F, Cl, Br, I, Me, Et, CF₃, cyclopropyl, cyano, OMe, OEt, CH₂OH, CH₂OMe and OH; the remainder of R^(AA), R^(BB), R^(CC) and R^(DD), (if present) are H; R^(C1), R^(C3) and R^(C4) are independently selected from H, Cl, F, Br, Me, OMe, OEt, cyano, CF₃, CH₂OH, CH₂OMe, C₂₋₄ alkenyl and C₅heterocyclyl; R^(C11), R^(C13) and R^(C14) are independently selected from H, Cl, F, Br, Me, OMe, OEt, cyano, CF₃, CH₂OH, CH₂OMe, C₂₋₄alkenyl and C₅heterocyclyl.

A second aspect of the present invention provides a compound of the first aspect for use in a method of therapy. The second aspect also provides a pharmaceutical composition comprising a compound of the first aspect and a pharmaceutically acceptable excipient.

A third aspect of the present invention provides a method of treatment or prevention of a disease ameliorated by the modulation of STING, comprising administering to a patient in need of treatment, a compound of the first aspect of the invention or a pharmaceutical composition of the second aspect of the invention. The third aspect of the present invention also provides the use of a compound of the first aspect of the invention in the manufacture of a medicament for treating or preventing disease ameliorated by the modulation of STING, and a compound of the first aspect of the invention or pharmaceutical composition thereof for use in the treatment or preventing of disease ameliorated by the modulation of STING.

Definitions

C₃₋₆ Cycloalkyl: The term “C₃₋₆ cycloalkyl” as used herein, pertains to a monovalent moiety obtained by removing a hydrogen atom from a carbon atom of a saturated cyclic hydrocarbon compound having from 3 to 6 carbon atoms. Examples of C₃₋₆ cycloalkyl groups include, but are not limited to, cyclopropyl (C₃), cyclobutyl (C₄), cyclopentyl (C₅) and cyclohexyl (C₆).

C₃₋₇Heterocyclyl: The term “C₃₋₇ heterocyclyl” as used herein, pertains to a monovalent moiety obtained by removing a hydrogen atom from a ring atom of a monocyclic heterocyclic compound, which moiety has from 3 to 7 ring atoms; of which from 1 to 2 atoms are heteroatoms, chosen from oxygen, sulfur or nitrogen. The C₃₋₇heterocyclyl groups may be non-aromatic or aromatic ring systems. Aromatic C₃₋₇heterocyclyl groups may be referred to as C₃₋₇heteroaryl groups.

In this context, the prefixes (e.g. C₃₋₇) denote the number of ring atoms, or range of number of ring atoms, whether carbon atoms or heteroatoms. Thus a prefix may in some instances be interchanged with an alternative prefix defining the number of ring member atoms, for example the prefix “O₃₋₇” may be interchanged with the prefix “3- to 7-membered”. In some embodiments, the C₃₋₇ heterocyclyl moieties in the compounds of the invention may be C₃, C₄, C₅, C₆ or C₇ heterocyclyls or any combination of these different sized rings/ring systems, such as O₃₋₆, C₄₋₇ or C₅₋₆ heterocyclyl groups.

Examples of C₃₋₇ heterocyclyl groups include, but are not limited to, those derived from:

N₁: aziridine (C₃), azetidine (C₄), pyrrolidine (tetrahydropyrrole) (C₅), pyrroline (e.g., 3-pyrroline, 2,5-dihydropyrrole) (C₅), 2H-pyrrole or 3H-pyrrole (isopyrrole, isoazole) (C₅), pyrrole (C₅), piperidine (C₆), dihydropyridine (C₆), tetrahydropyridine (C₆), pyridine (C₆), azepine (C₇), azepane (C₇); N₂: diazirine (O₃) diazetidine (C₄), imidazolidine (C₅), pyrazolidine (diazolidine) (C₅), imidazoline (C₅), pyrazoline (dihydropyrazole) (C₅), imidazole (C₅), pyrazole (C₅), piperazine (C₆), pyrazine (C₆), pyrimidine (C₆), pyridazine (C₆), diazepine (C₇), diazepane (C₇); O₁: oxetane (C₄), tetrahydrofuran (C₅); oxane (C₆); O₂: dioxetane (C₄), dioxolane (C₅); dioxane (C₆), dioxole (C₅); N₁O₁: tetrahydrooxazole (C₅), dihydrooxazole (C₅), tetrahydroisoxazole (C₅), dihydroisoxazole (C₅), isoxazole (C₅), oxazole (C₅), morpholine (C₆), tetrahydrooxazine (O₆), dihydrooxazine (C₆), oxazine (C₆); S₁: thiirane (C₃), thietane (C₄), thiolane (tetrahydrothiophene) (C₅), thiophene (C₅), thiane (tetrahydrothiopyran) (C₆), thiepane (C₇); N₁S₁: thiazoline (C₅), thiazolidine (C₅), thiazole (C₅), isothiazole (C₅), thiomorpholine (C₆), thiazine (C₆); O₁S₁: oxathiolidene (C₅), isoxthiolidine (C₅), oxathiole (C₅), isoxathiole (C₅) and oxathiane (thioxane) (C₆).

C₁₋₄ Alkyl: The term “C₁₋₄ alkyl” as used herein, pertains to a monovalent moiety obtained by removing a hydrogen atom from a carbon atom of a saturated hydrocarbon compound having from 1 to 4 carbon atoms.

Examples of saturated alkyl groups include, but are not limited to, Me: methyl (C₁), Et: ethyl (C₂), Pr: propyl (C₃), and Bu: butyl (C₄).

Examples of saturated linear alkyl groups include, but are not limited to, methyl (C₁), ethyl (C₂), nPr: n-propyl (C₃), and nBu: n-butyl (C₄).

Examples of saturated branched alkyl groups include, but are not limited to, iPr: iso-propyl (C₃, —C(CH₃)₂), iBu: iso-butyl (C₄), sBu: sec-butyl (C₄) and tBu: tert-butyl (C₄).

C₂₋₄ Alkenyl: The term “C₂₋₄ alkenyl” as used herein, pertains to an alkyl group having from 2 to 4 carbon atoms and having one or more carbon-carbon double bonds.

Examples of unsaturated alkenyl groups include, but are not limited to, ethenyl (vinyl, —CH═CH₂), 1-propenyl (—CH═CH—CH₃), 2-propenyl (allyl, —CH—CH═CH₂), isopropenyl (1-methylvinyl, —C(CH₃)═CH₂) and butenyl (C₄).

C₁₋₄ fluoroalkyl: The term “C₁₋₄ fluoroalkyl” as used herein, pertains to a C₁₋₄ alkyl group, substituted with one or more fluorine atoms.

Alkoxy: —OR wherein R is a C₁₋₄ alkyl group as defined above. It can be represented as —O—C₁₋₄ alkyl. Examples of alkoxy groups include, but are not limited to, methoxy (OMe, C), ethoxy (OEt, C₂), propyloxy (C₃), and butyloxy (C₄).

Alkyl carbamoyl: —NHC(═O)OR wherein R is a C₁₋₄ alkyl group as defined above. Examples of alkyl carbamoyl groups include, but are not limited to, —N(H)C(═O)OCH₃, —N(H)C(═O)OCH₂CH₃, and —N(H)C(═O)OC(CH₃)₃.

Alkyl carbamoyl ester: —OC(═O)NRR′ wherein R and R′ are independently selected from H and C₁₋₄ alkyl as defined above. Examples of alkyl carbamoyl ester groups include, but are not limited to, —OC(═O)N(CH₃)₂, and —OC(═O)N(H)CH₃.

Alkyl carboxyl ester: —OC(═O)OR wherein R is a C₁₋₄ alkyl group as defined above. Examples of alkyl carboxyl ester groups include, but are not limited to, —OC(═O)OCH₃, —OC(═O)OCH₂CH₃, —OC(═O)OC(CH₃)₃, and —OC(═O)OCH(CH₃)₂.

Amino: —N(R)R′ wherein R and R′ are independently selected from H and C₁₋₄ alkyl as defined above. Examples of an amino group include, but are not limited to, —NH₂, —N(H)CH₃, —N(H)C(CH₃)₂, —N(CH₃)₂, —N(CH₂CH₃)₂.

Amido (carbamoyl, carbamyl, aminocarbonyl, carboxamide, aminoacyl):

—C(═O)N(R)R′ wherein R and R′ are independently selected from H and C₁₋₄ alkyl as defined above. Examples of an amido group include, but are not limited to, C(═O)NH₂, —C(═O)N(H)CH₃, —C(═O)N(CH₃)₂, —C(═O)N(H)CH₂CH₃, and —C(═O)N(CH₂CH₃)₂.

Acylamido: —N(R)C(═O)R′ wherein R and R′ are independently selected from H and C₁₋₄ alkyl as defined above. Examples of an acylamido group include, but are not limited to, —N(H)C(═O)CH₂CH₃, —N(H)C(═O)CH₃ and —N(CH₃)C(═O)CH₃.

Phenyl: —C₆H₅, wherein the phenyl may itself be optionally substituted by one or more C₁₋₄alkyl groups, one or more C₁₋₄ fluoroalkyl groups, one or more C₁₋₄ alkoxy groups, one or more halo substituents and one or more cyano substituents.

Benzyl: —CH₂-phenyl, wherein phenyl is as defined above.

Ester (carboxylate, carboxylic acid ester, oxycarbonyl): —C(═O)OR, wherein R is an ester substituent, for example, a C₁₋₄ alkyl group, a C₃₋₇ heterocyclyl group, or a phenyl group, as defined above, preferably a C₁₋₄ alkyl group. Examples of ester groups include, but are not limited to, —C(═O)OCH₃, —C(═O)OCH₂CH₃, —C(═O)OC(CH₃)₃, and —C(═O)OPh.

Acyloxy (reverse ester): —OC(═O)R, wherein R is an acyloxy substituent, for example, a C₁₋₄alkyl group, a C₃₋₇ heterocyclyl group, or a phenyl group, as defined above, preferably a C₁₋₄alkyl group. Examples of acyloxy groups include, but are not limited to, —OC(═O)CH₃ (acetoxy), —OC(═O)CH₂CH₃, —OC(═O)C(CH₃)₃ and —OC(═O)Ph. Further examples of acyloxy groups include, but are not limited to, methylester (C₁), ethylester (C₂), propylester (C₃) and butylester (C₄).

Naturally occurring amino acid: The term “a naturally occurring amino acid”, as used herein, pertains to a monovalent moiety obtained by removing a hydrogen atom from a carboxyl group or an amino group on one of the amino acid compounds found commonly in nature (for example, alanine, arginine, asparagine, aspartate, cysteine, glycine, glutamine, glutamate, histidine, isoleucine, leucine, lysine, methionine, phenylalanine, proline, serine, threonine, tryptophan, tyrosine and valine). The amino acid is particularly selected from isoleucine, leucine and valine, most particularly valine.

In each of these groups the carbon atom which is bonded to both a carboxyl and an amino group is known as the α carbon and the carboxyl and amino groups to which it is attached are the α-carboxyl and α-amino groups. Naturally occurring amino acids are optionally substituted with a protecting group on the α-amino group or any other amino group on the moiety, protecting groups include but are not limited to acetyl, methyl, fluorenylmethoxycarbonyl (Fmoc), carboxybenzyl (Cbz; benzyloxycarbonyl), phthalimido and tertbutyl carbamate (boc) groups.

Phosphonate ester: —P(O)(OR)OR′, wherein R and R′ are independently selected from C₁₋₄alkyl as defined above. Examples of a phosphonate ester include, but are not limited to —P(O)(OEt)₂.

Cyano: —C≡N.

Pivaloyloxymethyl: A group of formula

Tetrazolyl: A group of formula

Oxo-oxadiazolyl: A 5 membered heterocyclic compound have two nitrogen ring atoms, an oxygen ring atom, and an oxo substituent. Examples of oxo-oxadiazolyl groups include:

5-oxo-4H-1,2,4-oxadiazol-3-yl 2-oxo-3H-1,3,4-oxadiazol-5-yl

2H-Triazol-4-yl: A group of formula:

As used herein, except where the context requires otherwise, the term “comprise” and variations of the term, such as “comprising”, “comprises” and “comprised”, are not intended to exclude further additives, components, integers or steps.

It must be noted that as used herein and in the appended claims, the singular forms “a”, “an” and “the” include plural reference unless the context clearly dictates otherwise. Thus, for example, a reference to “a salt” may include a plurality of salts and a reference to “at least one heteroatom” may include one or more heteroatoms, and so forth.

The term “and/or” can mean “and” or “or”.

The term “(s)” following a noun contemplates the singular or plural form, or both.

Various features of the invention are described with reference to a certain value, or range of values. These values are intended to relate to the results of the various appropriate measurement techniques, and therefore should be interpreted as including a margin of error inherent in any particular measurement technique. Some of the values referred to herein are denoted by the term “about” to at least in part account for this variability. The term “about”, when used to describe a value, may mean an amount within ±10%, ±5%, ±1% or ±0.1% of that value.

Includes Other Forms

Unless otherwise specified, included in the above are the well known ionic, salt, solvate, and protected forms of these substituents. For example, a reference to carboxylic acid (—COOH) also includes the anionic (carboxylate) form (—COO⁻), a salt or solvate thereof, as well as conventional protected forms. Similarly, a reference to an amino group includes the protonated form (—N⁺HR¹R²), a salt or solvate of the amino group, for example, a hydrochloride salt, as well as conventional protected forms of an amino group. Similarly, a reference to a hydroxyl group also includes the anionic form (—O⁻), a salt or solvate thereof, as well as conventional protected forms.

Salts

It may be convenient or desirable to prepare, purify, and/or handle a corresponding salt of the active compound, for example, a pharmaceutically-acceptable salt. Examples of pharmaceutically acceptable salts are discussed in Berge, et al., J. Pharm. Sci., 66, 1-19 (1977).

For example, if the compound is anionic, or has a functional group which may be anionic (e.g. —COOH may be —COO⁻), then a salt may be formed with a suitable cation. Examples of suitable inorganic cations include, but are not limited to, alkali metal ions such as Na⁺ and K⁺, alkaline earth cations such as Ca²⁺ and Mg²⁺, and other cations such as Al⁺³. Examples of suitable organic cations include, but are not limited to, ammonium ion (i.e. NH₄+) and substituted ammonium ions (e.g. NH₃R⁺, NH₂R₂ ⁺, NHR₃ ⁺, NR₄ ⁺). Examples of some suitable substituted ammonium ions are those derived from: ethylamine, diethylamine, dicyclohexylamine, triethylamine, butylamine, ethylenediamine, ethanolamine, diethanolamine, piperazine, benzylamine, phenylbenzylamine, choline, meglumine, and tromethamine, as well as amino acids, such as lysine and arginine. An example of a common quaternary ammonium ion is N(CH₃)₄ ⁺.

If the compound is cationic, or has a functional group which may be cationic (e.g. —NH₂ may be —NH₃ ⁺), then a salt may be formed with a suitable anion. Examples of suitable inorganic anions include, but are not limited to, those derived from the following inorganic acids: hydrochloric, hydrobromic, hydroiodic, sulfuric, sulfurous, nitric, nitrous, phosphoric, and phosphorous.

Examples of suitable organic anions include, but are not limited to, those derived from the following organic acids: 2-acetyoxybenzoic, acetic, ascorbic, aspartic, benzoic, camphorsulfonic, cinnamic, citric, edetic, ethanedisulfonic, ethanesulfonic, fumaric, glucoheptonic, gluconic, glutamic, glycolic, hydroxymaleic, hydroxynaphthalene carboxylic, isethionic, lactic, lactobionic, lauric, maleic, malic, methanesulfonic, mucic, oleic, oxalic, palmitic, pamoic, pantothenic, phenylacetic, phenylsulfonic, propionic, pyruvic, salicylic, stearic, succinic, sulfanilic, tartaric, toluenesulfonic, trifluoroacetic acid and valeric. Examples of suitable polymeric organic anions include, but are not limited to, those derived from the following polymeric acids: tannic acid, carboxymethyl cellulose.

Solvates

It may be convenient or desirable to prepare, purify, and/or handle a corresponding solvate of the active compound. The term “solvate” is used herein in the conventional sense to refer to a complex of solute (e.g. active compound, salt of active compound) and solvent. If the solvent is water, the solvate may be conveniently referred to as a hydrate, for example, a mono-hydrate, a di-hydrate, a tri-hydrate, etc.

Prodrugs

The active compounds may be prepared as prodrugs. In particular, the carboxy groups R¹ and R¹¹ may be present as ester, amido (—C(═O)NH₂) and substituted amido groups. In some embodiments R¹ and R¹¹ may be independently selected from —C(═O)OR^(P), —C(═O)NH₂ and —C(═O)NHR^(P), where R^(P) is selected from:

-   -   (i) C₃₋₆cycloalkyl;     -   (ii) C₃₋₇ heterocyclyl optionally substituted with a group         selected from:         -   methyl; and         -   ester; and     -   (iii) linear or branched C₁₋₄alkyl optionally substituted with a         group selected from:         -   alkoxy;         -   amino;         -   amido;         -   acylamido;         -   acyloxy;         -   alkyl carboxyl ester;         -   alkyl carbamoyl;         -   alkyl carbamoyl ester;         -   phenyl;         -   phosphonate ester;         -   C₃₋₇heterocyclyl optionally substituted with a group             selected from methyl and oxo; and         -   a naturally occurring amino acid, optionally N-substituted             with a group         -   selected from methyl, acetyl and boc;

In some embodiments, R¹ and R¹¹ are —C(═O)NH₂. In these embodiments, the compounds are of formula Ic:

where A¹A⁴, A¹¹-A¹⁴, R^(C1), R^(C3), R^(C4), R^(C11), R^(C13), R^(C14) and Y are as defined above.

In some embodiments R^(P) is optionally substituted linear or branched C₁₋₄alkyl. In some embodiments R^(P) is unsubstituted C₁₋₄alkyl. In some embodiments R^(P) is substituted C₁₋₄alkyl.

When R^(P) is C₁₋₄ alkyl, in some of these embodiments R^(P) is methyl. In other of these embodiments, R^(P) is ethyl. In other of these embodiments, R^(P) is propyl (e.g. iso-propyl, n-propyl). In other of these embodiments, R^(P) is butyl (e.g. iso-butyl, sec-butyl, tert-butyl).

In some embodiments, R^(P) is C₃₋₆cycloalkyl. In some of these embodiments, R^(P) is cyclopropyl. In other of these embodiments, R^(P) is cyclobutyl. In other of these embodiments, R^(P) is cyclopentyl. In other of these embodiments, R^(P) is cyclohexyl.

In some embodiments, R^(P) is C₃₋₇heterocyclyl. In some of these embodiments, the C₃₋₇heterocyclyl has a single nitrogen ring atom. In some of these embodiments, R^(P) is azetidinyl, pyrrolidinyl or piperidinyl. In some of these embodiments, R^(P) is azetidinyl. In some of these embodiments, R^(P) is piperidinyl.

Substituents on R^(P)

In some embodiments, when R^(P) is C₃₋₇heterocyclyl, it is substituted with a group selected from methyl and ester. In some embodiments, when R^(P) is C₃₋₇heterocyclyl, it is substituted with methyl. In some embodiments, when R^(P) is C₃₋₇heterocyclyl, it is substituted with ester.

In some embodiments, when R^(P) is linear or branched C₁₋₄alkyl, it is substituted with a group selected from alkoxy, amino, amido, acylamido, acyloxy, alkyl carboxyl ester, alkyl carbamoyl, alkyl carbamoyl ester, phenyl, phosphonate ester C₃₋₇heterocyclyl optionally substituted with group selected from methyl and oxo, and a naturally occurring amino acid, optionally N-substituted with a group selected from methyl, acetyl and boc.

In some embodiments, when R^(P) is linear or branched C₁₋₄alkyl, it is substituted with a group selected from acyloxy and phenyl. In some embodiments, when R^(P) is linear or branched C₁₋₄alkyl substituted with acyloxy, it is pivaloyloxymethyl; a group of formula:

In some embodiments, when R^(P) is linear or branched C₁₋₄alkyl substituted with acyloxy, it is propanoyloxyisobutyl; a group of formula:

In some embodiments, when R^(P) is linear or branched C₁₋₄alkyl substituted with phenyl, it is benzyl.

In some embodiments, when R^(P) is linear or branched C₁₋₄alkyl, it is substituted with a naturally occurring amino acid, optionally N-substituted with a group selected from methyl, acetyl and boc. In some embodiments when R^(P) is linear or branched C₁₋₄alkyl substituted with a naturally occurring amino acid, the naturally occurring amino acid is valine. In some embodiments when R^(P) is linear or branched C₁₋₄alkyl substituted with a naturally occurring amino acid, the naturally occurring amino acid is N-methyl valine. In some embodiments when R^(P) is linear or branched C₁₋₄alkyl substituted with a naturally occurring amino acid, the naturally occurring amino acid is N-acetyl valine. In some embodiments when R^(P) is linear or branched C₁₋₄alkyl substituted with a naturally occurring amino acid, the naturally occurring amino acid is N-boc valine.

In some embodiments, when R^(P) is linear or branched C₁₋₄alkyl, it is substituted with amino. In some embodiments, when R^(P) is linear or branched C₁₋₄alkyl, it is substituted with amido. In some embodiments, when R^(P) is linear or branched C₁₋₄alkyl, it is substituted with acylamido. In some embodiments, when R^(P) is linear or branched C₁₋₄alkyl, it is substituted with acyloxy. In some embodiments, when R^(P) is linear or branched C₁₋₄alkyl, it is substituted with alkyl carboxyl ester. In some embodiments, when R^(P) is linear or branched C₁₋₄alkyl, it is substituted with alkyl carbamoyl. In some embodiments, when R^(P) is linear or branched C₁₋₄alkyl, it is substituted with alkyl carbamoyl ester. In some embodiments, when R^(P) is linear or branched C₁₋₄alkyl, it is substituted with phenyl. In some embodiments, when R^(P) is linear or branched C₁₋₄alkyl, it is substituted with phosphonate ester.

In some embodiments, when R^(P) is linear or branched C₁₋₄alkyl, it is substituted with C₃₋₇heterocyclyl, optionally substituted with a group selected from methyl and oxo. In some embodiments when R^(P) is linear or branched C₁₋₄alkyl substituted with C₃₋₇heterocyclyl, the C₃₋₇heterocyclyl is dioxole, optionally substituted with a group selected from methyl and oxo.

Isomers

Certain compounds of the invention may exist in one or more particular geometric, optical, enantiomeric, diasteriomeric, epimeric, atropic, stereoisomeric, tautomeric, conformational, or anomeric forms, including but not limited to, cis- and trans-forms; E- and Z-forms; c-, t-, and r-forms; endo- and exo-forms; R-, S-, and meso-forms; D- and L-forms; d- and l-forms; (+) and (−) forms; keto-, enol-, and enolate-forms; syn- and anti-forms; synclinal- and anticlinal-forms; α- and β-forms; axial and equatorial forms; boat-, chair-, twist-, envelope-, and halfchair-forms; M- and P-forms; Δ- and ∧-forms; R_(a)- and S_(a)-forms; and combinations thereof, hereinafter collectively referred to as “isomers” (or “isomeric forms”).

The term “chiral” refers to molecules which have the property of non-superimposability of the mirror image partner, while the term “achiral” refers to molecules which are superimposable on their mirror image partner.

The term “stereoisomers” refers to compounds which have identical chemical constitution, but differ with regard to the arrangement of the atoms or groups in space.

“Diastereomer” refers to a stereoisomer with two or more centers of chirality and whose molecules are not mirror images of one another. Diastereomers have different physical properties, e.g. melting points, boiling points, spectral properties, and reactivities. Mixtures of diastereomers may separate under high resolution analytical procedures such as electrophoresis and chromatography.

“Enantiomers” refer to two stereoisomers of a compound which are non-superimposable mirror images of one another.

“Atropisomers” refer to two stereoisomers of a compound which arise due to restricted rotation about a single bond, wherein rotation about said bond is sufficiently hindered that individual conformers can be isolated and the conformers are either non-superimposable mirror images of one another (enantiomers) or stereoisomers that are not mirror images of one another (diastereomers).

Stereochemical definitions and conventions used herein generally follow S. P. Parker, Ed., McGraw-Hill Dictionary of Chemical Terms (1984) McGraw-Hill Book Company, New York; and Eliel, E. and Wilen, S., “Stereochemistry of Organic Compounds”, John Wiley & Sons, Inc., New York, 1994. The compounds of the invention may contain asymmetric or chiral centers, and therefore exist in different stereoisomeric forms. It is intended that all stereoisomeric forms of the compounds of the invention, including but not limited to, diastereomers, enantiomers and atropisomers, as well as mixtures thereof such as racemic mixtures, form part of the present invention. Many organic compounds exist in optically active forms, i.e., they have the ability to rotate the plane of plane-polarized light. In describing an optically active compound, the prefixes D and L, or R and S, M and P, Δ and ∧, R_(a) and S_(a) are used to denote the absolute configuration of the molecule about its chiral center(s). The prefixes d and I or (+) and (−) are employed to designate the sign of rotation of plane-polarized light by the compound, with (−) or l meaning that the compound is levorotatory. A compound prefixed with (+) or d is dextrorotatory. For a given chemical structure, a (+)-stereoisomer and a (−)-stereoisomer are identical except that they are non-superimposable mirror images of one another. Each of these specific stereoisomers may be referred to as an enantiomer, and a mixture of such isomers is often called an enantiomeric mixture. A 50:50 mixture of enantiomers is referred to as a racemic mixture or a racemate, which may occur where there has been no stereoselection or stereospecificity in a chemical reaction or process. The terms “racemic mixture” and “racemate” refer to an equimolar mixture of two enantiomeric species, devoid of optical activity.

Of particular relevance to the compounds of the present invention is atropisomerism, where there may be restricted rotation about the single bond between the two aromatic ring systems in each half of the dimer—this may occur when A1 and/or A11 are of a certain size.

Note that, except as discussed below for tautomeric forms, specifically excluded from the term “isomers”, as used herein, are structural (or constitutional) isomers (i.e. isomers which differ in the connections between atoms rather than merely by the position of atoms in space). For example, a reference to a methoxy group, —OCH₃, is not to be construed as a reference to its structural isomer, a hydroxymethyl group, —CH₂OH. Similarly, a reference to ortho-chlorophenyl is not to be construed as a reference to its structural isomer, meta-chlorophenyl. However, a reference to a class of structures may well include structurally isomeric forms falling within that class (e.g. C₁₋₇ alkyl includes n-propyl and iso-propyl; butyl includes n-, iso-, sec-, and tert-butyl; methoxyphenyl includes ortho-, meta-, and para-methoxyphenyl).

The above exclusion does not pertain to tautomeric forms, for example, keto-, enol-, and enolate-forms, as in, for example, the following tautomeric pairs: keto/enol (illustrated below), imine/enamine, amide/imino alcohol, amidine/amidine, nitroso/oxime, thioketone/enethiol, N-nitroso/hyroxyazo, and nitro/aci-nitro.

The term “tautomer” or “tautomeric form” refers to structural isomers of different energies which are interconvertible via a low energy barrier. For example, proton tautomers (also known as prototropic tautomers) include interconversions via migration of a proton, such as keto-enol and imine-enamine isomerizations. Valence tautomers include interconversions by reorganization of some of the bonding electrons.

Note that specifically included in the term “isomer” are compounds with one or more isotopic substitutions. For example, H may be in any isotopic form, including ¹H, ²H (D), and ³H (T); C may be in any isotopic form, including ¹²C, ¹³C, and ¹⁴C; O may be in any isotopic form, including ¹⁶O and ¹⁸O; and the like.

Examples of isotopes that can be incorporated into compounds of the invention include isotopes of hydrogen, carbon, nitrogen, oxygen, phosphorous, fluorine, and chlorine, such as, but not limited to ²H (deuterium, D), ³H (tritium), ¹¹C, ¹³C, ¹⁴C, ¹⁵N, ¹⁸F, ³¹P, ³²P, ³⁵S, ³⁶Cl, and ¹²⁵I. Various isotopically labeled compounds of the present invention, for example those into which radioactive isotopes such as ³H, ¹³C, and ¹⁴C are incorporated. Such isotopically labelled compounds may be useful in metabolic studies, reaction kinetic studies, detection or imaging techniques, such as positron emission tomography (PET) or single-photon emission computed tomography (SPECT) including drug or substrate tissue distribution assays, or in radioactive treatment of patients. Deuterium labelled or substituted therapeutic compounds of the invention may have improved DMPK (drug metabolism and pharmacokinetics) properties, relating to distribution, metabolism, and excretion (ADME). Substitution with heavier isotopes such as deuterium may afford certain therapeutic advantages resulting from greater metabolic stability, for example increased in vivo half-life or reduced dosage requirements. An ¹⁸F labeled compound may be useful for PET or SPECT studies. Isotopically labeled compounds of this invention and prodrugs thereof can generally be prepared by carrying out the procedures disclosed in the schemes or in the examples and preparations described below by substituting a readily available isotopically labeled reagent for a non-isotopically labeled reagent. Further, substitution with heavier isotopes, particularly deuterium (i.e., ²H or D) may afford certain therapeutic advantages resulting from greater metabolic stability, for example increased in vivo half-life or reduced dosage requirements or an improvement in therapeutic index. It is understood that deuterium in this context is regarded as a substituent. The concentration of such a heavier isotope, specifically deuterium, may be defined by an isotopic enrichment factor. In the compounds of this invention any atom not specifically designated as a particular isotope is meant to represent any stable isotope of that atom.

Unless otherwise specified, a reference to a particular compound includes all such isomeric forms, including (wholly or partially) racemic and other mixtures thereof. Methods for the preparation (e.g. asymmetric synthesis) and separation (e.g. fractional crystallisation and chromatographic means) of such isomeric forms are either known in the art or are readily obtained by adapting the methods taught herein, or known methods, in a known manner.

Polymorphs

Compounds disclosed herein (in any of the various forms described herein) that form crystalline solids may demonstrate polymorphism. All polymorphic forms of the compounds, are within the scope of the disclosure.

N-Oxides

Nitrogen containing groups may also be oxidised to form an N-oxide.

Therapeutic Indications

Compounds disclosed herein may provide a therapeutic benefit in a number of disorders, in particular, in the treatment or prevention of diseases ameliorated by the modulation of STING.

One aspect of the invention provides methods of treatment or prevention of STING mediated diseases and disorders, in which agonizing STING is beneficial. Exemplary diseases/disorders includes, but are not limited to, cancer and infectious disease (such as those caused by viruses, e.g., HIV, HBV, HCV, HPV, influenza and coronaviruses (including rhinoviruses, SARS, MERS and SARS-CoV-2), and bacteria). Another aspect of the invention provides the use of a STING agonist as a vaccine adjuvant.

In one embodiment, this invention provides a compound of the invention for use in therapy. This invention also provides a compound of Formula I, or a pharmaceutically acceptable salt thereof, for use in therapy. This invention particularly provides a compound of Formula I, or a pharmaceutically acceptable salt thereof, for use in the treatment of a STING-mediated disease or disorder.

This invention also provides a compound of Formula I, or a pharmaceutically acceptable salt thereof, for use as a vaccine adjuvant. There is also therefore provided an immugenic composition or vaccine adjuvant comprising a compound of Formula I, or a pharmaceutically acceptable salt thereof. In a further embodiment of the invention, there is provided a composition comprising a compound of Formula I, or a pharmaceutically acceptable salt thereof, and one or more immunostimulatory agents.

In another embodiment, this invention provides a compound of the invention for use in the treatment of a STING-mediated disease or disorder and/or for use as an immugenic composition or a vaccine adjuvant. In another embodiment, this invention provides a compound of Formula I, or a pharmaceutically acceptable salt thereof, for use in the amelioration of organ injury or damage sustained as a result of a STING-mediated disease or disorder.

The invention further provides for the use of a compound of the invention in the manufacture of a medicament for treatment of a STING-mediated disease or disorder. The invention further provides for the use of a compound of Formula I, or a salt thereof, particularly a pharmaceutically acceptable salt thereof, in the manufacture of a medicament for treatment of a STING-mediated disease or disorder, for example the diseases and disorders recited herein.

The invention further provides for the use of a compound of Formula I, or a salt thereof, particularly a pharmaceutically acceptable salt thereof, in the manufacture of a vaccine.

There is further provided the use of a compound of Formula I, or a pharmaceutically acceptable salt thereof, for the manufacture of an immugenic composition comprising an antigen or antigenic composition, for the treatment or prevention of disease. There is further provided the use of a compound of Formula I, or a pharmaceutically acceptable salt thereof, for the manufacture of a vaccine composition comprising an antigen or antigenic composition, for the treatment or prevention of disease.

In another embodiment, the invention is directed to a method of treating a STING-mediated disease or disorder comprising administering a therapeutically effective amount of a compound of this invention to a human in need thereof. In another embodiment, the invention is directed to a method of treating a STING-mediated disease or disorder comprising administering a therapeutically effective amount of a compound of Formula I or a salt, particularly a pharmaceutically acceptable salt thereof, to a human in need thereof.

Infectious Diseases

The compounds of this invention may be used to treat an infectious disease, which is any disease instigated by or coincident with an infection from a pathogen. Pathogens are broadly defined as any species of organism that is foreign to a human tissue environment. Common disease causing pathogens include bacteria (many like TB), viruses (many like HBV, HIV, influenza, coronaviruses) and parasitic protozoans (like P falciparum that causes malaria). The compounds of this invention may be used to treat infectious diseases derived from bacteria, such as TB infection Mycobacterium tuberculosis), Chlamydia, Tularemia infection Francisella tularensis), Plasmodium infection or infections from DNA or RNA virus. The compounds of this invention may be used to treat infectious diseases derived from the DNA virus families: Herpesviridae (herpes simplex virus-1, Kaposi's sarcoma-associated virus and Epstein-Barr virus), Papillomaviridae (human papilloma virus), Adenovirus and Hepadnaviridae (Hepatitis B virus). Examples of RNA virus families include Retroviridae (human immunodeficiency virus) Flaviviridae (Dengue virus, Hepatitis C virus), Orthomyxoviridae (influenza), and Coronaviridae (human coronavirus and SARS coronavirus, including SARS-CoV-2).

Cancer

As used herein, the terms “cancer”, “neoplasm,” and “tumor” are used interchangeably and, in either the singular or plural form, refer to cells that have undergone a malignant transformation that makes them pathological to the host organism. Primary cancer cells can be readily distinguished from non-cancerous cells by well-established techniques, particularly histological examination. The definition of a cancer cell, as used herein, includes not only a primary cancer cell, but any cell derived from a cancer cell ancestor. This includes metastasized cancer cells, and in vitro cultures and cell lines derived from cancer cells. When referring to a type of cancer that normally manifests as a solid tumor, a “clinically detectable” tumor is one that is detectable on the basis of tumor mass; e.g., by procedures such as computed tomography (CT) scan, magnetic resonance imaging (MRI), X-ray, ultrasound or palpation on physical examination, and/or which is detectable because of the expression of one or more cancer-specific antigens in a sample obtainable from a patient. Tumors may be a hematopoietic (or hematologic or hematological or blood-related) cancer, for example, cancers derived from blood cells or immune cells, which may be referred to as “liquid tumors.” Specific examples of clinical conditions based on hematologic tumors include leukemias, such as chronic myelocytic leukemia, acute myelocytic leukemia, chronic lymphocytic leukemia and acute lymphocytic leukemia; plasma cell malignancies such as multiple myeloma, MGUS and Waldenstrom's macroglobulinemia; lymphomas such as non-Hodgkin's lymphoma, Hodgkin's lymphoma; and the like.

The cancer may be any cancer in which an abnormal number of blast cells or unwanted cell proliferation is present or that is diagnosed as a hematological cancer, including both lymphoid and myeloid malignancies. Myeloid malignancies include, but are not limited to, acute myeloid (or myelocytic or myelogenous or myeloblasts) leukemia (undifferentiated or differentiated), acute promyeloid (or promyelocytic or promyelogenous or promyeloblastic) leukemia, acute myelomonocytic (or myelomonoblastic) leukemia, acute monocytic (or monoblastic) leukemia, erythroleukemia and megakaryocytic (or megakaryoblastic) leukemia. These leukemias may be referred together as acute myeloid (or myelocytic or myelogenous) leukemia (AML). Myeloid malignancies also include myeloproliferative disorders (MPD) which include, but are not limited to, chronic myelogenous (or myeloid) leukemia (CML), chronic myelomonocytic leukemia (CMML), essential thrombocythemia (or thrombocytosis), and polycythemia vera (PCV). Myeloid malignancies also include myelodysplasia (or myelodysplastic syndrome or MDS), which may be referred to as refractory anemia (RA), refractory anemia with excess blasts (RAEB), and refractory anemia with excess blasts in transformation (RAEBT); as well as myelofibrosis (MFS) with or without agnogenic myeloid metaplasia.

Hematopoietic cancers also include lymphoid malignancies, which may affect the lymph nodes, spleens, bone marrow, peripheral blood, and/or extranodal sites. Lymphoid cancers include B-cell malignancies, which include, but are not limited to, B-cell non-Hodgkin's lymphomas (B-NHLs). B-NHLs may be indolent (or low-grade), intermediate-grade (or aggressive) or high-grade (very aggressive). Indolent B cell lymphomas include follicular lymphoma (FL); small lymphocytic lymphoma (SLL); marginal zone lymphoma (MZL) including nodal MZL, extranodal MZL, splenic MZL and splenic MZL with villous lymphocytes; lymphoplasmacytic lymphoma (LPL); and mucosa-associated-lymphoid tissue (MALT or extranodal marginal zone) lymphoma. Intermediate-grade B-NHLs include mantle cell lymphoma (MCL) with or without leukemic involvement, diffuse large cell lymphoma (DLBCL), follicular large cell (or grade 3 or grade 3B) lymphoma, and primary mediastinal lymphoma (PML). High-grade B-NHLs include Burkitt's lymphoma (BL), Burkitt-like lymphoma, small non-cleaved cell lymphoma (SNCCL) and lymphoblastic lymphoma. Other B-NHLs include immunoblastic lymphoma (or immunocytoma), primary effusion lymphoma, HIV associated (or AIDS related) lymphomas, and post-transplant lymphoproliferative disorder (PTLD) or lymphoma. B-cell malignancies also include, but are not limited to, chronic lymphocytic leukemia (CLL), prolymphocytic leukemia (PLL), Waldenstrom's macroglobulinemia (WM), hairy cell leukemia (HCL), large granular lymphocyte (LGL) leukemia, acute lymphoid (or lymphocytic or lymphoblastic) leukemia, and Castleman's disease. NHL may also include T-cell non-Hodgkin's lymphoma s(T-NHLs), which include, but are not limited to T-cell non-Hodgkin's lymphoma not otherwise specified (NOS), peripheral T-cell lymphoma (PTCL), anaplastic large cell lymphoma (ALCL), angioimmunoblastic lymphoid disorder (AILD), nasal natural killer (NK) cell/T-cell lymphoma, gamma/delta lymphoma, cutaneous T cell lymphoma, mycosis fungoides, and Sezary syndrome.

Hematopoietic cancers also include Hodgkin's lymphoma (or disease) including classical Hodgkin's lymphoma, nodular sclerosing Hodgkin's lymphoma, mixed cellularity Hodgkin's lymphoma, lymphocyte predominant (LP) Hodgkin's lymphoma, nodular LP Hodgkin's lymphoma, and lymphocyte depleted Hodgkin's lymphoma. Hematopoietic cancers also include plasma cell diseases or cancers such as multiple myeloma (MM) including smoldering MM, monoclonal gammopathy of undetermined (or unknown or unclear) significance (MGUS), plasmacytoma (bone, extra medullary), lymphoplasmacytic lymphoma (LPL), Waldenstrom's Macroglobulinemia, plasma cell leukemia, and primary amyloidosis (AL). Hematopoietic cancers may also include other cancers of additional hematopoietic cells, including polymorphonuclear leukocytes (or neutrophils), basophils, eosinophils, dendritic cells, platelets, erythrocytes and natural killer cells. Tissues which include hematopoietic cells referred herein to as “hematopoietic cell tissues” include bone marrow; peripheral blood; thymus; and peripheral lymphoid tissues, such as spleen, lymph nodes, lymphoid tissues associated with mucosa (such as the gut-associated lymphoid tissues), tonsils, Peyer's patches and appendix, and lymphoid tissues associated with other mucosa, for example, the bronchial linings.

Examples of cancer diseases and conditions in which a compounds of this invention may have potentially beneficial antitumor effects include, but are not limited to, cancers of the lung, bone, pancreas, skin, head, neck, uterus, ovaries, stomach, colon, breast, esophagus, small intestine, bowel, endocrine system, thyroid gland, parathyroid gland, adrenal gland, urethra, prostate, penis, testes, ureter, bladder, kidney or liver; rectal cancer; cancer of the anal region; carcinomas of the fallopian tubes, endometrium, cervix, vagina, vulva, renal pelvis, renal cell; sarcoma of soft tissue; myxoma; rhabdomyoma; fibroma; lipoma; teratoma; cholangiocarcinoma; hepatoblastoma; angiosarcoma; hemangioma; hepatoma; fibrosarcoma; chondrosarcoma; myeloma; chronic or acute leukemia; lymphocytic lymphomas; primary CNS lymphoma; neoplasms of the CNS; spinal axis tumors; squamous cell carcinomas; synovial sarcoma; malignant pleural mesotheliomas; brain stem glioma; pituitary adenoma; bronchial adenoma; chondromatous hamartoma; mesothelioma; Hodgkin's Disease or a combination of one or more of the foregoing cancers. Suitably the present invention relates to a method for treating or lessening the severity of cancers selected from the group consisting of brain (gliomas), glioblastomas, astrocytomas, glioblastoma multiforme, Bannayan-Zonana syndrome, Cowden disease, Lhermitte-Duclos disease, Wilm's tumor, Ewing's sarcoma, Rhabdomyosarcoma, ependymoma, medulloblastoma, head and neck, kidney, liver, melanoma, ovarian, pancreatic, adenocarcinoma, ductal adenocarcinoma, adenosquamous carcinoma, acinar cell carcinoma, glucagonoma, insulinoma, prostate, sarcoma, osteosarcoma, giant cell tumor of bone, thyroid, lymphoblastic T cell leukemia, chronic myelogenous leukemia, chronic lymphocytic leukemia, hairy-cell leukemia, acute lymphoblastic leukemia, acute myelogenous leukemia, chronic neutrophilic leukemia, acute lymphoblastic T cell leukemia, plasmacytoma, immunoblastic large cell leukemia, mantle cell leukemia, multiple myeloma, megakaryoblastic leukemia, multiple myeloma, acute megakaryocytic leukemia, promyelocytic leukemia, erythroleukemia, malignant lymphoma, Hodgkin's lymphoma, non-Hodgkin's lymphoma, lymphoblastic T cell lymphoma, Burkitt's lymphoma, follicular lymphoma, neuroblastoma, bladder cancer, urothelial cancer, vulval cancer, cervical cancer, endometrial cancer, renal cancer, mesothelioma, esophageal cancer, salivary gland cancer, hepatocellular cancer, gastric cancer, nasopharangeal cancer, buccal cancer, cancer of the mouth, GIST (gastrointestinal stromal tumor) and testicular cancer.

In some embodiments, the compounds of the present invention may be used to treat solid or liquid tumors. In some embodiments, the compounds of the present invention may be used to treat sarcoma, breast cancer, colorectal cancer, gastroesophageal cancer, melanoma, non-small cell lung cancer (NSCLC), clear cell renal cell carcinoma (RCC), lymphomas, squamous cell carcinoma of the head and neck (SCCHN), hepatocellular carcinoma (HCC), and/or Non Hodgkin lymphoma (NHL). Suitably the present invention relates to a method for treating or lessening the severity of pre-cancerous syndromes in a mammal, including a human, wherein the pre-cancerous syndrome is selected from: cervical intraepithelial neoplasia, monoclonal gammopathy of unknown significance (MGUS), myelodysplastic syndrome, aplastic anemia, cervical lesions, skin nevi (pre-melanoma), prostatic intraepithelial (intraductal) neoplasia (PIN), Ductal Carcinoma in situ (DCIS), colon polyps and severe hepatitis or cirrhosis.

In one aspect the human has a solid tumor. In one aspect the tumor is selected from head and neck cancer, gastric cancer, melanoma, renal cell carcinoma (RCC), esophageal cancer, non-small cell lung carcinoma, prostate cancer, colorectal cancer, ovarian cancer and pancreatic cancer. In one aspect the human has one or more of the following: colorectal cancer (CRC), esophageal, cervical, bladder, breast, head and neck, ovarian, melanoma, renal cell carcinoma (RCC), EC squamous cell, non-small cell lung carcinoma, mesothelioma, and prostate cancer. In another aspect the human has a liquid tumor such as diffuse large B cell lymphoma (DLBCL), multiple myeloma, chronic lymphoblastic leukemia (CLL), follicular lymphoma, acute myeloid leukemia and chronic myelogenous leukemia. In one embodiment, the compounds of the present invention may be useful for treatment of skin cancers (e.g., non-melanoma skin cancer, squamous cell carcinoma, basal cell carcinoma) or actinic keratosis. In addition to a field effect for clearing superficial skin cancers, the compounds of the present invention may prevent the development of subsequent skin cancers and pre-malignant actinic keratosis in treated patients.

Autoimmune Diseases

Autoimmune diseases associated include, but are not limited to STING associated vasculitis with onset at infancy (SAVI), Aicardi Goutieres syndrome (AGS), chilblain lupus, ataxia telanogiectasia (also referred to as Louis-Bar Syndrome), retinal vasculopathy with cerebral leukodystrophy (RCVL), systemic lupus erythematosus (SLE), cutaneous lupus, lupus nephritis, psoriasis, diabetes mellitus including insulin-dependent diabetes mellitus (IDDM), dermatomyositis, human immunodeficiency virus (HIV), AIDS, polymyositis, systemic sclerosis (scleroderma), and Sjogren's syndrome (SS), rheumatoid arthritis, psoriatic arthritis, polyarthritis, myasthenia gravis, polyarteritis nodosa, vasculitis, cutaneous vasculitis, anti-neutrophil cytoplasmic antibody (ANCA)-associated vasculitis, Henoch-Schonlein purpura, autoimmune hepatitis, primary sclerosing cholangitis, Wegener's granulomatosis, microscopic polyangiitis, Behcet's disease, spondylitis, giant cell arteritis, polymyalgia rheumatic, Raynaud's phenomenon, primary biliary cirrhosis, primary angiitis of the central nervous system microscopic polyangiitis, neuromyelitis optica and mixed connective tissue disease.

Inflammation

Inflammation represents a group of vascular, cellular and neurological responses to trauma. Inflammation can be characterized as the movement of inflammatory cells such as monocytes, neutrophils and granulocytes into the tissues. This is usually associated with reduced endothelial barrier function and oedema into the tissues. Inflammation can be classified as either acute or chronic. Acute inflammation is the initial response of the body to harmful stimuli and is achieved by the increased movement of plasma and leukocytes from the blood into the injured tissues. A cascade of biochemical event propagates and matures the inflammatory response, involving the local vascular system, the immune system, and various cells within the injured tissue. Prolonged inflammation, known as chronic inflammation, leads to a progressive shift in the type of cells which are present at the site of inflammation and is characterized by simultaneous destruction and healing of the tissue from the inflammatory process.

When occurring as part of an immune response to infection or as an acute response to trauma, inflammation can be beneficial and is normally self-limiting. However, inflammation can be detrimental under various conditions. This includes the production of excessive inflammation in response to infectious agents, which can lead to significant organ damage and death (for example, in the setting of sepsis). Moreover, chronic inflammation is generally deleterious and is at the root of numerous chronic diseases, causing severe and irreversible damage to tissues. In such settings, the immune response is often directed against self-tissues (autoimmunity), although chronic responses to foreign entities can also lead to bystander damage to self-tissues.

The aim of anti-inflammatory therapy is therefore to reduce this inflammation, to inhibit autoimmunity when present, and to allow for the physiological process or healing and tissue repair to progress.

The compounds of this invention may be used to treat inflammation of any tissue and organs of the body, including musculoskeletal inflammation, vascular inflammation, neural inflammation, digestive system inflammation, ocular inflammation, inflammation of the reproductive system, and other inflammation, as exemplified below.

Musculoskeletal inflammation refers to any inflammatory condition of the musculoskeletal system, particularly those conditions affecting skeletal joints, including joints of the hand, wrist, elbow, shoulder, jaw, spine, neck, hip, knee, ankle, and foot, and conditions affecting tissues connecting muscles to bones such as tendons. Examples of musculoskeletal inflammation which may be treated with compounds of the invention include arthritis (including, for example, osteoarthritis, rheumatoid arthritis, psoriatic arthritis, ankylosing spondylitis, acute and chronic infectious arthritis, arthritis associated with gout and pseudogout, and juvenile idiopathic arthritis), tendonitis, synovitis, tenosynovitis, bursitis, fibrositis (fibromyalgia), epicondylitis, myositis, and osteitis (including, for example, Paget's disease, osteitis pubis, and osteitis fibrosa cystic).

Ocular inflammation refers to inflammation of any structure of the eye, including the eye lids. Examples of ocular inflammation which may be treated with the compounds of the invention include blepharitis, blepharochalasis, conjunctivitis, dacryoadenitis, keratitis, keratoconjunctivitis sicca (dry eye), scleritis, trichiasis, and uveitis.

Examples of inflammation of the nervous system which may be treated with the compounds of the invention include encephalitis, Guillain-Barre syndrome, meningitis, neuromyotonia, narcolepsy, multiple sclerosis, myelitis, CNS vasculitis, and schizophrenia.

Examples of inflammation of the vasculature or lymphatic system which may be treated with the compounds of the invention include arthrosclerosis, arthritis, phlebitis, vasculitis, and lymphangitis.

Examples of inflammatory conditions of the digestive system which may be treated with the compounds of the invention include cholangitis, cholecystitis, enteritis, enterocolitis, gastritis, gastroenteritis, inflammatory bowel disease (such as Crohn's disease and ulcerative colitis), ileitis, and proctitis.

Examples of inflammatory conditions of the reproductive system which may be treated with the compounds of the invention include cervicitis, chorioamnionitis, endometritis, epididymitis, omphalitis, oophoritis, orchitis, salpingitis, tubo-ovarian abscess, urethritis, vaginitis, vulvitis, and vulvodynia.

The compounds of this invention may be used to treat autoimmune conditions having an inflammatory component. Such conditions include acute disseminated alopecia universalise, Behcet's disease, Chagas' disease, STING associated vasculitis with onset at infancy (SAVI), Aicardi Goutieres syndrome (AGS), chilblain lupus, ataxia telangiectasia (also referred to as Louis-Bar Syndrome), retinal vasculopathy with cerebral leukodystrophy (RCVL), ANCA)-associated vasculitis, chronic fatigue syndrome, dysautonomia, encephalomyelitis, ankylosing spondylitis, aplastic anemia, hidradenitis suppurativa, autoimmune hepatitis, autoimmune oophoritis, celiac disease, Crohn's disease, diabetes mellitus type 1, giant cell arteritis, goodpasture's syndrome, Grave's disease, Guillain-Barre syndrome, Hashimoto's disease, Henoch-Schonlein purpura, Kawasaki's disease, lupus erythematosus, microscopic colitis, microscopic polyarteritis, mixed connective tissue disease, multiple sclerosis, myasthenia gravis, opsoclonus myoclonus syndrome, optic neuritis, ord's thyroiditis, pemphigus, polyarteritis nodosa, polymyalgia, rheumatoid arthritis, Reiter's syndrome, Sjogren's syndrome, temporal arteritis, Wegener's granulomatosis, warm autoimmune hemolytic anemia, interstitial cystitis, lyme disease, morphea, psoriasis, sarcoidosis, scleroderma, ulcerative colitis, and vitiligo.

The compounds of this invention may be used to treat T-cell mediated hypersensitivity diseases having an inflammatory component. Such conditions include contact hypersensitivity, contact dermatitis (including that due to poison ivy), uticaria, skin allergies, respiratory allergies (hayfever, allergic rhinitis) and gluten-sensitive enteropathy (Celiac disease).

Other inflammatory conditions which may be treated with the compounds of this invention include, for example, appendicitis, dermatitis, dermatomyositis, endocarditis, fibrositis, gingivitis, glossitis, hepatitis, hidradenitis suppurativa, iritis, laryngitis, mastitis, myocarditis, nephritis, otitis, pancreatitis, parotitis, pericarditis, peritonitis, pharyngitis, pleuritis, pneumonitis, prostatitis, pyelonephritis, and stomatitis, transplant rejection (involving organs such as kidney, liver, heart, lung, pancreas (e.g., islet cells), bone marrow, cornea, small bowel, skin allografts, skin homografts, and heart valve xenografts, serum sickness, and graft vs host disease), acute pancreatitis, chronic pancreatitis, acute respiratory distress syndrome, Sezary syndrome, congenital adrenal hyperplasia, nonsuppurative thyroiditis, hypercalcemia associated with cancer, pemphigus, bullous dermatitis herpetiformis, severe erythema multiforme, exfoliative dermatitis, seborrheic dermatitis, seasonal or perennial allergic rhinitis, bronchial asthma, contact dermatitis, atopic dermatitis, drug hypersensitivity reactions, allergic conjunctivitis, keratitis, herpes zoster ophthalmicus, iritis and iridocyclitis, chorioretinitis, optic neuritis, symptomatic sarcoidosis, fulminating or disseminated pulmonary tuberculosis chemotherapy, idiopathic thrombocytopenic purpura in adults, secondary thrombocytopenia in adults, acquired (autoimmune) hemolytic anemia, leukemia and lymphomas in adults, acute leukemia of childhood, regional enteritis, autoimmune vasculitis, multiple sclerosis, chronic obstructive pulmonary disease, solid organ transplant rejection, sepsis. Preferred treatments include treatment of transplant rejection, rheumatoid arthritis, psoriatic arthritis, multiple sclerosis, Type 1 diabetes, asthma, inflammatory bowel disease, systemic lupus erythematosus, psoriasis, chronic pulmonary disease, and inflammation accompanying infectious conditions (e.g., sepsis). In one embodiment, the compounds of this invention may be used to treat asthma.

Cellular Proliferation

The compounds of the present invention may also be useful in the treatment of one or more diseases afflicting mammals which are characterized by cellular proliferation in the area of disorders associated with neo-vascularization and/or vascular permeability including blood vessel proliferative disorders including arthritis (rheumatoid arthritis) and restenosis; fibrotic disorders including hepatic cirrhosis and atherosclerosis; mesangial cell proliferative disorders include glomerulonephritis, diabetic nephropathy, malignant nephrosclerosis, thrombotic microangiopathy syndromes, proliferative retinopathies, organ transplant rejection and glomerulopathies; and metabolic disorders include psoriasis, diabetes mellitus, chronic wound healing, inflammation and neurodegenerative diseases.

Neurodegenerative Diseases

The compounds of this invention may be used to treat neurodegenerative diseases. Exemplary neurodegenerative diseases includes, but are not limited to, multiple sclerosis, Huntington's disease, Alzheimer's disease, Parkinson's disease, amyotrophic lateral sclerosis (ALS).

Combinations

The compounds of this invention may be employed alone or in combination with other therapeutic agents. As modulators of the immune response, the compounds of this invention may also be used in monotherapy or used in combination with another therapeutic agent in the treatment of diseases and conditions in which modulation of STING is beneficial. Combination therapies according to the present invention thus comprise the administration of a compound of Formula I or a pharmaceutically acceptable salt thereof, and at least one other therapeutically active agent. In one embodiment, combination therapies according to the present invention comprise the administration of at least one compound of Formula I or a pharmaceutically acceptable salt thereof, and at least one other therapeutic agent. The compound(s) of Formula I and pharmaceutically acceptable salts thereof, and the other therapeutic agent(s) may be administered together in a single pharmaceutical composition or separately and, when administered separately this may occur simultaneously or sequentially in any order. The amounts of the compound(s) of Formula I and pharmaceutically acceptable salts thereof, and the other therapeutic agent(s) and the relative timings of administration will be selected in order to achieve the desired combined therapeutic effect. Thus in a further aspect, there is provided a combination comprising a compound of Formula I, or a pharmaceutically acceptable salt thereof, together with one or more other therapeutic agents.

The compounds of Formula I and pharmaceutically acceptable salts thereof may be used in combination with one or more other therapeutic agents which may be useful in the prevention or treatment of allergic disease, inflammatory disease, or autoimmune disease, for example; antigen immunotherapy, anti-histamines, steroids, NSAIDs, bronchodilators (e.g. beta 2 agonists, adrenergic agonists, anticholinergic agents, theophylline), methotrexate, leukotriene modulators and similar agents; monoclonal antibody therapy such as anti-IgE, anti-TNF, anti-IL-5, anti-IL-6, anti-IL-12, anti-IL-1 and similar agents; receptor therapies e.g. etanercept and similar agents; antigen non-specific immunotherapies (e.g. interferon or other cytokines/chemokines, cytokine/chemokine receptor modulators, cytokine agonists or antagonists, TLR agonists and similar agents).

The compounds of Formula I and pharmaceutically acceptable salts thereof may be used in combination with radiotherapy and/or surgery and/or at least one other therapeutic agent which may be useful in the treatment of cancer and pre-cancerous syndromes. Any anti-neoplastic agent that has activity versus a susceptible tumor being treated may be utilized in the combination. Typical anti-neoplastic agents useful include, but are not limited to, (a) anti-microtubule agents such as diterpenoids (e.g. paclitaxel, docetaxel) and vinca alkaloids (e.g. vinblastine, vincristine, and vinorelbine); (b) platinum coordination complexes (e.g. oxaliplatin, cisplatin and carboplatin); (c) alkylating agents such as nitrogen mustards (e.g. cyclophosphamide, melphalan, and chlorambucil), oxazaphosphorines, alkylsulfonates (e.g. busulfan), nitrosoureas (e.g. carmustine), and triazenes (e.g. dacarbazine); (d) antibiotic agents such as anthracyclins (e.g. daunorubicin and doxorubicin), actinomycins (e.g. dactinomycin) and bleomycins; (e) topoisomerase II inhibitors such as epipodophyllotoxins (e.g. etoposide and teniposide); (f) antimetabolites such as purine and pyrimidine analogues and anti-folate compounds (e.g. fluorouracil, methotrexate, cytarabine, mercaptopurine, thioguanine, and gemcitabine); (g) topoisomerase I inhibitors such as camptothecins (e.g. irinotecan, topotecan, and various optical forms of 7-(4-methylpiperazino-methylene)-10,11-ethylenedioxy-20-camptothecin); (h) hormones and hormonal analogues (e.g. adrenocorticosteroids such as prednisone and prednisolone which are useful in the treatment of malignant lymphoma and acute leukemia in children; aminoglutethimide and other aromatase inhibitors such as anastrozole, letrozole, vorozole, and exemestane useful in the treatment of adrenocortical carcinoma and hormone dependent breast carcinoma containing estrogen receptors; progestins such as megestrol acetate useful in the treatment of hormone dependent breast cancer and endometrial carcinoma; estrogens, and anti-estrogens such as fulvestrant, flutamide, nilutamide, bicalutamide, cyproterone acetate and 5-reductases such as finasteride and dutasteride, useful in the treatment of prostatic carcinoma and benign prostatic hypertrophy; anti-estrogens such as tamoxifen, toremifene, raloxifene, droloxifene, iodoxyfene, as well as selective estrogen receptor modulators (SERMS) such those described in U.S. Pat. Nos. 5,681,835, 5,877,219, and 6,207,716, useful in the treatment of hormone dependent breast carcinoma and other susceptible cancers; and gonadotropin-releasing hormone (GnRH) and analogues thereof which stimulate the release of leutinizing hormone (LH) and/or follicle stimulating hormone (FSH) for the treatment prostatic carcinoma, for instance, LHRH agonists and antagonists such as goserelin acetate and leuprolide); (i) signal transduction pathway inhibitors; (j) non-receptor tyrosine angiogenesis inhibitors; (k) immunotherapeutic agents (e.g. ex-vivo and in-vivo approaches to increase the immunogenicity of patient tumor cells, such as transfection with cytokines such as interleukin 2, interleukin 4 or granulocyte-macrophage colony stimulating factor, approaches to decrease T-cell energy, approaches using transfected immune cells such as cytokine-transfected dendritic cells, approaches using cytokine-transfected tumor cell lines and approaches using anti-idiotypic antibodies); (I) proapoptotic agents (e.g. bcl-2 antisense oligonucleotides); (m) cell cycle signalling inhibitors; (n) immuno-oncology agents and (o) immunostimulatory agents.

Signal Transduction Pathway Inhibitors

Signal transduction pathway inhibitors are those inhibitors, which block or inhibit a chemical process which evokes an intracellular change. As used herein this change is cell proliferation or differentiation. Signal transduction inhibitors useful in the present invention include inhibitors of receptor tyrosine kinases, non-receptor tyrosine kinases, SH2/SH3 domain blockers, serine/threonine kinases, phosphotidyl inositol-3 kinases, myo-inositol signalling, and Ras oncogenes.

Several protein tyrosine kinases catalyze the phosphorylation of specific tyrosyl residues in various proteins involved in the regulation of cell growth. Such protein tyrosine kinases can be broadly classified as receptor or non-receptor kinases.

Receptor tyrosine kinases are transmembrane proteins having an extracellular ligand binding domain, a transmembrane domain, and a tyrosine kinase domain. Receptor tyrosine kinases are involved in the regulation of cell growth and are generally termed growth factor receptors. Inappropriate or uncontrolled activation of many of these kinases, i.e. aberrant kinase growth factor receptor activity, for example by over-expression or mutation, has been shown to result in uncontrolled cell growth. Accordingly, the aberrant activity of such kinases has been linked to malignant tissue growth. Consequently, inhibitors of such kinases could provide cancer treatment methods. Growth factor receptors include, for example, epidermal growth factor receptor (EGFr), platelet derived growth factor receptor (PDGFr), erbB2, erbB4, ret, vascular endothelial growth factor receptor (VEGFr), tyrosine kinase with immunoglobulin-like and epidermal growth factor homology domains (TIE-2), insulin growth factor-I (IGFI) receptor, macrophage colony stimulating factor (cfms), BTK, ckit, cmet, fibroblast growth factor (FGF) receptors, Trk receptors (TrkA, TrkB, and TrkC), ephrin (eph) receptors, and the RET protooncogene. Several inhibitors of growth receptors are under development and include ligand antagonists, antibodies, tyrosine kinase inhibitors and anti-sense oligonucleotides. Growth factor receptors and agents that inhibit growth factor receptor function are described, for instance, in Kath 2000; Shawver 1997; and Lofts 1994.

Tyrosine kinases, which are not growth factor receptor kinases, are termed nonreceptor tyrosine kinases. Non-receptor tyrosine kinases useful in the present invention, which are targets or potential targets of anti-cancer drugs, include cSrc, Lck, Fyn, Yes, Jak, cAbl, FAK (Focal adhesion kinase), Brutons tyrosine kinase, and Bcr-Abl. Such non-receptor kinases and agents which inhibit non-receptor tyrosine kinase function are described in Sinh 1999; and Bolen 1997.

SH2/SH3 domain blockers are agents that disrupt SH2 or SH3 domain binding in a variety of enzymes or adaptor proteins including, PI3-K p85 subunit, Src family kinases, adaptor molecules (She, Crk, Nek, Grb2) and Ras-GAP. SH2/SH3 domains as targets for anticancer drugs are discussed in Smithgall 1995.

Inhibitors of Serine/Threonine Kinases including MAP kinase cascade blockers which include blockers of Raf kinases (rafk), Mitogen or Extracellular Regulated Kinase (MEKs), and Extracellular Regulated Kinases (ERKs); and Protein kinase C family member blockers including blockers of PKCs (alpha, beta, gamma, epsilon, mu, lambda, iota, zeta). IkB kinase family (IKKa, IKKb), PKB family kinases, akt kinase family members, and TGF beta receptor kinases. Such Serine/Threonine kinases and inhibitors thereof are described in Yamamoto 1999; Brodt 2000; Massague 1996; Philip 1995; Lackey 2000; U.S. Pat. No. 6,268,391; and Martinez-Lacaci 2000.

Inhibitors of Phosphotidyl inositol-3 Kinase family members including blockers of Pekinese, ATM, DNA-PK, and Ku are also useful in the present invention. Such kinases are discussed in Abraham 1996; Canman 1998; Jackson 1997; and Zhong 2000.

Also useful in the present invention are Myo-inositol signalling inhibitors such as phospholipase C blockers and Myoinositol analogues. Such signal inhibitors are described in Powis 1994.

Another group of signal transduction pathway inhibitors are inhibitors of Ras Oncogene. Such inhibitors include inhibitors of farnesyltransferase, geranyl-geranyl transferase, and CAAX proteases as well as anti-sense oligonucleotides, ribozymes and immunotherapy. Such inhibitors have been shown to block ras activation in cells containing wild type mutant ras, thereby acting as antiproliferation agents. Ras oncogene inhibition is discussed in Scharovsky 2000; Ashby 1998; and Oliff 1999.

As mentioned above, antibody antagonists to receptor kinase ligand binding may also serve as signal transduction inhibitors. This group of signal transduction pathway inhibitors includes the use of humanized antibodies to the extracellular ligand binding domain of receptor tyrosine kinases. For example Imclone C225 EGFR specific antibody (see Green 2000); Herceptin® erbB2 antibody (see Stern 2000); and 2CB VEGFR2 specific antibody (see Brekken 2000).

Non-Receptor Tyrosine Angiogenesis Inhibitors

Anti-angiogenic therapeutic agents including non-receptor MEK angiogenesis inhibitors may also be useful. Anti-angiogenic agents such as those which inhibit the effects of vascular endothelial growth factor, (for example the anti-vascular endothelial cell growth factor antibody bevacizumab [Avastin™], and compounds that work by other mechanisms (for example linomide, inhibitors of integrin av133 function, endostatin and angiostatin).

Cell Cycle Signalling Inhibitors

Cell cycle signalling inhibitors inhibit molecules involved in the control of the cell cycle. A family of protein kinases called cyclin dependent kinases (CDKs) and their interaction with a family of proteins termed cyclins controls progression through the eukaryotic cell cycle. The coordinate activation and inactivation of different cyclin/CDK complexes is necessary for normal progression through the cell cycle. Several inhibitors of cell cycle signalling are under development. For instance, examples of cyclin dependent kinases, including CDK2, CDK4, and CDK6 and inhibitors for the same are described in, for instance, Rosania 2000.

Immuno-Modulators

Additional examples of other therapeutic agents (e.g., anti-neoplastic agent) for use in combination or co-administered with a compound of Formula I are immuno-modulators. As used herein “immuno-modulators” refer to any substance including monoclonal antibodies that affects the immune system. Immuno-modulators can be used as anti-neoplastic agents for the treatment of cancer. For example, immune-modulators include, but are not limited to, anti-CTLA-4 antibodies such as ipilimumab (YERVOY) and anti-PD-1 antibodies (Opdivo/nivolumab and Keytruda/pembrolizumab). Other immuno-modulators include, but are not limited to, ICOS antibodies, OX-40 antibodies, PD-L1 antibodies, LAGS antibodies, TIM-3 antibodies, 41 BB antibodies and GITR antibodies.

Anti-PD-L1 Agents

Additional examples of other therapeutic agents (anti-neoplastic agent) for use in combination or co-administered with a compound of this invention are anti-PD-L1 agents. Anti-PD-L1 antibodies and methods of making the same are known in the art. Such antibodies to PD-L1 may be polyclonal or monoclonal, and/or recombinant, and/or humanized. Exemplary PD-L1 antibodies are disclosed in U.S. Pat. Nos. 8,217,149, 8,383,796, 8,552,154, 9,212,224, and 8,779,108, and US Patent Appln. Pub. Nos. 2011/0280877, 2014/0341902 and 2013/0045201. Additional exemplary antibodies to PD-L1 (also referred to as CD274 or B7-H1) and methods for use are disclosed in U.S. Pat. Nos. 7,943,743, 8,168,179, and 7,595,048; WO2014/055897, WO2016/007235; and US Patent Appln. Pub. Nos. 2013/0034559 and 2015/0274835. PD-L1 antibodies are in development as immunomodulatory agents for the treatment of cancer.

In one embodiment, the antibody to PD-L1 is an antibody disclosed in U.S. Pat. No. 8,217,149. In another embodiment, the anti-PD-L1 antibody comprises the CDRs of an antibody disclosed in U.S. Pat. No. 8,217,149. In another embodiment, the antibody to PD-L1 is an antibody disclosed in U.S. Pat. No. 8,779,108. In another embodiment, the anti-PD-L1 antibody comprises the CDRs of an antibody disclosed in U.S. Pat. No. 8,779,108. In another embodiment, the antibody to PD-L1 is an antibody disclosed in US Patent Appln. Pub. No. 2013/0045201. In another embodiment, the anti-PD-L1 antibody comprises the CDRs of an antibody disclosed in US Patent Appln. Pub. No. 2013/0045201. In one embodiment, the anti-PD-L1 antibody is BMS-936559 (MDX-1105), which was described in WO 2007/005874. In another embodiment, the anti-PD-L1 antibody is MPDL3280A (RG7446). In another embodiment, the anti-PD-L1 antibody is MED14736, which is an anti-PD-L1 monoclonal antibody described in WO 2011/066389 and US 2013/034559. In another embodiment, the anti-PD-L1 antibody is TECENTRIQ™ (atezolizumab), which is an anti-PD-L1 cancer immunotherapy which was approved in the US in May 2016 for specific types of bladder cancer. In another embodiment, anti-PD-L1 antibody is YW243.55.570 which is an anti-PD-L1 described in WO 2010/077634 and U.S. Pat. No. 8,217,149. Examples of anti-PD-L1 antibodies useful for the methods of this invention, and methods for making thereof are described in PCT patent application WO 2010/077634, WO 2007/005874, WO 2011/066389, U.S. Pat. No. 8,217,149, and US 2013/034559.

PD-1 Antagonist

Additional examples of other therapeutic agents (anti-neoplastic agent) for use in combination or co-administered with a compound of this invention are PD-1 antagonists. “PD-1 antagonist” means any chemical compound or biological molecule that blocks binding of PD-L1 expressed on a cancer cell to PD-1 expressed on an immune cell (T cell, B cell or NKT cell) and preferably also blocks binding of PD-L2 expressed on a cancer cell to the immune-cell expressed PD-1. Alternative names or synonyms for PD-1 and its ligands include: PDCDI, PDI, CD279 and SLEB2 for PD-1; PDCDILI, PDLI, B7H1, B7-4, CD274 and B7-H for PD-LI; and PDCD1 L2, PDL2, B7-DC, Btdc and CD273 for PD-L2. In any embodiments of the aspects or embodiments of the present invention in which a human individual is to be treated, the PD-1 antagonist blocks binding of human PD-L1 to human PD-1, and preferably blocks binding of both human PD-L1 and PD-L2 to human PD-1. Human PD-1 amino acid sequences can be found in NCBI Locus No.: NP_005009. Human PD-L1 and PD-L2 amino acid sequences can be found in NCBI Locus No.: NP_054862 and NP_079515, respectively.

PD-1 antagonists useful in any of the aspects of the present invention include a monoclonal antibody (mAb), or antigen binding fragment thereof, which specifically binds to PD-1 or PD-L1, and preferably specifically binds to human PD-1 or human PD-L1. The mAb may be a human antibody, a humanized antibody or a chimeric antibody, and may include a human constant region. In some embodiments, the human constant region is selected from the group consisting of IgGI, IgG2, IgG3 and IgG4 constant regions, and in preferred embodiments, the human constant region is an IgGI or IgG4 constant region. In some embodiments, the antigen binding fragment is selected from the group consisting of Fab, Fab′-SH, F(ab′)2, scFv and Fv fragments.

Examples of mAbs that bind to human PD-1, and useful in the various aspects and embodiments of the present invention, are described in U.S. Pat. Nos. 7,488,802, 7,521,051, 8,008,449, 8,354,509, 8,168,757, WO2004/004771, WO2004/072286, WO2004/056875, and US2011/0271358. Specific anti-human PD-1 mAbs useful as the PD-1 antagonist in any of the aspects and embodiments of the present invention include: MK-3475, a humanized IgG4 mAb with the structure described in WHO Drug Information, Vol. 27, No. 2, pages 161-162 (2013) and which comprises the heavy and light chain amino acid sequences shown in FIG. 6; nivolumab, a human IgG4 mAb with the structure described in WHO Drug Information, Vol. 27, No. 1, pages 68-69 (2013) and which comprises the heavy and light chain amino acid sequences shown in FIG. 7; the humanized antibodies h409A11, h409A16 and h409A17, which are described in WO2008/156712, and AMP-514, which is being developed by Medimmune.

Other PD-1 antagonists useful in the any of the aspects and embodiments of the present invention include an immunoadhesin that specifically binds to PD-1, and preferably specifically binds to human PD-1, e.g., a fusion protein containing the extracellular or PD-1 binding portion of PD-L1 or PD-L2 fused to a constant region such as an Fc region of an immunoglobulin molecule. Examples of immunoadhesion molecules that specifically bind to PD-1 are described in WO2010/027827 and WO2011/066342. Specific fusion proteins useful as the PD-1 antagonist in the treatment method, medicaments and uses of the present invention include AMP-224 (also known as B7-DCIg), which is a PD-L2-FC fusion protein and binds to human PD-1.

Other examples of mAbs that bind to human PD-L1, and useful in the treatment method, medicaments and uses of the present invention, are described in WO2013/019906, WO2010/077634 and U.S. Pat. No. 8,383,796. Specific anti-human PD-L1 mAbs useful as the PD-1 antagonist in the treatment method, medicaments and uses of the present invention include MPDL3280A, BMS-936559, MED14736, MSB00107180.

KEYTRUDA/pembrolizumab is an anti-PD-1 antibody marketed for the treatment of lung cancer by Merck. The amino acid sequence of pembrolizumab and methods of using are disclosed in U.S. Pat. No. 8,168,757.

Opdivo/nivolumab is a fully human monoclonal antibody marketed by Bristol Myers Squibb directed against the negative immunoregulatory human cell surface receptor PD-1 (programmed death-1 or programmed cell death-I/POD-1) with immunopotentiation activity. Nivolumab binds to and blocks the activation of PD-1, an Ig superfamily transmembrane protein, by its ligands PD-L1 and PD-L2, resulting in the activation of T-cells and cell-mediated immune responses against tumor cells or pathogens. Activated PD-1 negatively regulates T-cell activation and effector function through the suppression of PI3K/Akt pathway activation. Other names for nivolumab include: BMS-936558, MDX-1106, and ONO-4538. The amino acid sequence for nivolumab and methods of using and making are disclosed in U.S. Pat. No. 8,008,449.

Antibodies to ICOS

Additional examples of other therapeutic agents (anti-neoplastic agent) for use in combination or co-administered with a compound of Formula I are antibodies to ICOS. ICOS is a co-stimulatory T cell receptor with structural and functional relation to the CD28/CTLA-4-Ig superfamily (Hutloff 1999). Activation of ICOS occurs through binding by ICOS-L (B7RP-1/B7-H2). Neither B7-1 nor B7-2 (ligands for CD28 and CTLA4) bind or activate ICOS. However, ICOS-L has been shown to bind weakly to both CD28 and CTLA-4 (Yao 2011). Expression of ICOS appears to be restricted to T cells. ICOS expression levels vary between different T cell subsets and on T cell activation status. ICOS expression has been shown on resting TH17, T follicular helper (TFH) and regulatory T (Treg) cells; however, unlike CD28; it is not highly expressed on naive THI and TH2 effector T cell populations (Paulos 2010). ICOS expression is highly induced on CD4+ and CD8+ effector T cells following activation through TCR engagement (Wakamatsu 2013).

CDRs for murine antibodies to human ICOS having agonist activity are shown in PCT/EP2012/055735 (WO 2012/131004). Antibodies to ICOS are also disclosed in WO 2008/137915, WO 2010/056804, EP1374902, EP1374901, and EP1125585. Agonist antibodies to ICOS or ICOS binding proteins are disclosed in WO2012/131004, WO 2014/033327, WO2016/120789, US20160215059, and US20160304610. In one embodiment, agonist antibodies to ICOS include ICOS binding proteins or antigen binding portions thereof comprising one or more of: CDRHI as set forth in SEQ ID NO:1; CDRH2 as set forth in SEQ ID NO:2; CDRH3 as set forth in SEQ ID NO:3; CDRLI as set forth in SEQ ID NO:4; CDRL2 as set forth in SEQ ID NO:5 and/or CDRL3 as set forth in SEQ ID NO:6 or a direct equivalent of each CDR wherein a direct equivalent has no more than two amino acid substitutions in said CDR as disclosed in WO2016/120789, which is incorporated by reference in its entirety herein. In one embodiment, the ICOS binding protein or antigen binding portion thereof is an agonist antibody to ICOS comprising a VH domain comprising an amino acid sequence at least 90% identical to the amino acid sequence set forth in SEQ ID NO:7 and/or a VL domain comprising an amino acid sequence at least 90% identical to the amino acid sequence as set forth in SEQ ID NO:8 as set forth in WO2016/120789 wherein said ICOS binding protein specifically binds to human ICOS. In one embodiment, the ICOS binding protein is an agonist antibody to ICOS comprising a VH domain comprising the amino acid sequence set forth in SEQ ID NO:7 and a VL domain comprising the amino acid sequence set forth in SEQ ID NO:8 as set forth in WO2016/120789. Yervoy (ipilimumab) is a fully human CTLA-4 antibody marketed by Bristol Myers Squibb. The protein structure of ipilimumab and methods are using are described in U.S. Pat. Nos. 6,984,720 and 7,605,238.

CD134, also known as OX40, is a member of the TNFR-superfamily of receptors which is not constitutively expressed on resting naive T cells, unlike CD28. OX40 is a secondary costimulatory molecule, expressed after 24 to 72 hours following activation; its ligand, OX40L, is also not expressed on resting antigen presenting cells, but is following their activation. Expression of OX40 is dependent on full activation of the T cell; without CD28, expression of OX40 is delayed and of fourfold lower levels. OX-40 antibodies, OX-40 fusion proteins and methods of using them are disclosed in U.S. Pat. Nos. 7,504,101; 7,758,852; 7,858,765; 7,550,140; 7,960,515; WO2012/027328; WO2013/028231. In one embodiment, the OX40 antigen binding protein is one disclosed in WO2012/027328 (PCT/US2011/048752), international filing date 23 Aug. 2011. In another embodiment, the antigen binding protein comprises the CDRs of an antibody disclosed in WO2012/027328 (PCT/US2011/048752), international filing date 23 Aug. 2011, or CDRs with 90% identity to the disclosed CDR sequences. In a further embodiment the antigen binding protein comprises a VH, a VL, or both of an antibody disclosed in WO2012/027328 (PCT/US2011/048752), international filing date 23 Aug. 2011, or a VH or a VL with 90% identity to the disclosed VH or VL sequences.

In another embodiment, the OX40 antigen binding protein is disclosed in WO2013/028231 (PCT/US2012/024570), international filing date 9 Feb. 2012, which is incorporated by reference in its entirety herein. In another embodiment, the antigen binding protein comprises the CDRs of an antibody disclosed in WO2013/028231 (PCT/US2012/024570), international filing date 9 Feb. 2012, or CDRs with 90% identity to the disclosed CDR sequences. In a further embodiment, the antigen binding protein comprises a VH, a VL, or both of an antibody disclosed in WO2013/028231 (PCT/US2012/024570), international filing date 9 Feb. 2012, or a VH or a VL with 90% identity to the disclosed VH or VL sequences. In one embodiment, the OX40 antigen binding protein is an isolated agonist antibody to OX40 comprising a light chain variable region having a sequence at least 90% identical to the amino acid sequence of SEQ ID NO: 10 as set forth in WO2013/028231 and a heavy chain variable region having a sequence at least 90% identical to the amino acid sequence of SEQ ID NO:4 as set forth in WO2013/028231. In one embodiment, the OX40 antigen binding protein is an isolated antibody comprising a light chain variable comprising the amino acid sequence of SEQ ID NO:10 as set forth in WO2013/028231 and a heavy chain variable region comprising the amino acid sequence of SEQ ID NO:4 as set forth in WO2013/028231.

Immunostimulatory Agent

Additional examples of other therapeutic agents for use in combination or coadministered with a compound of Formula I, or a salt thereof are immunostimulatory agents.

As used herein “immunostimulatory agent” refers to any agent that can stimulate the immune system. As used herein immunostimulatory agents include, but are not limited to, vaccine adjuvants, such as Toll-like receptor agonists, T-cell checkpoint blockers, such as mAbs to PD-1 and CTL4 and T-cell checkpoint agonist, such as agonist mAbs to OX-40 and ICOS. As used herein “immunostimulatory agent” refers to any agent that can stimulate the immune system. As used herein immunostimulatory agents include, but are not limited to, vaccine adjuvants.

The term “Toll-like receptor” (or “TLR”) as used herein refers to a member of the Toll-like receptor family of proteins or a fragment thereof that senses a microbial product and/or initiates an adaptive immune response. In one embodiment, a TLR activates a dendritic cell (DC). Toll-like receptors (TLRs) are a family of pattern recognition receptors that were initially identified as sensors of the innate immune system that recognize microbial pathogens. TLRs recognize distinct structures in microbes, often referred to as “PAMPs” (pathogen associated molecular patterns). Ligand binding to TLRs invokes a cascade of intra-cellular signalling pathways that induce the production of factors involved in inflammation and immunity. In humans, ten TLRs have been identified. TLRs that are expressed on the surface of cells include TLR-1, -2, -4, -5, and -6, while TLR-3, -7/8, and -9 are expressed with the ER compartment. Human DC subsets can be identified on the basis of distinct TLR expression patterns. By way of example, the myeloid or “conventional” subset of DC (mDC) expresses TLRs 1-8 when stimulated, and a cascade of activation markers (e.g. CD80, CD86, MHC class I and II, CCR7), pro-inflammatory cytokines, and chemokines are produced. A result of this stimulation and resulting expression is antigen-specific CD4+ and CD8+ T cell priming. These DCs acquire an enhanced capacity to take up antigens and present them in an appropriate form to T cells. In contrast, the plasmacytoid subset of DC (pDC) expresses only TLR7 and TLR9 upon activation, with a resulting activation of NK cells as well as T-cells. As dying tumor cells may adversely affect DC function, it has been suggested that activating DC with TLR agonists may be beneficial for priming anti-tumor immunity in an immunotherapy approach to the treatment of cancer. It has also been suggested that successful treatment of breast cancer using radiation and chemotherapy requires TLR4 activation.

TLR agonists known in the art and finding use in the present invention include, but are not limited to, the following: Pam3Cys, a TLRI/2 agonist; CFA, a TLR2 agonist; MALP2, a TLR2 agonist; Pam2Cys, a TLR2 agonist; FSL-I, a TLR-2 agonist; Hib-OMPC, a TLR-2 agonist; polyinosinic:polycytidylic acid (Poly I:C), a TLR3 agonist; polyadenosine-polyuridylic acid (poly AU), a TLR3 agonist; Polyinosinic-Polycytidylic acid stabilized with poly-L-lysine and carboxymethylcellulose (Hiltonol), a TLR3 agonist; bacterial flagellin a TLR5 agonist; imiquimod, a TLR7 agonist; resiquimod, a TLR7/8 agonist; loxoribine, a TLR7/8 agonist; and unmethylated CpG dinucleotide (CpG-ODN), a TLR9 agonist.

Additional TLR agonists known in the art and finding use in the present invention further include, but are not limited to aminoalkyl glucosaminide phosphates (AGPs) which bind to the TLR4 receptor are known to be useful as vaccine adjuvants and immunostimulatory agents for stimulating cytokine production, activating macrophages, promoting innate immune response, and augmenting antibody production in immunized animals. An example of a naturally occurring TLR4 agonist is bacterial LPS. An example of a semisynthetic TLR4 agonist is monophosphoryl lipid A (MPL). AGPs and their immunomodulating effects via TLR4 are disclosed in patent publications such as WO 2006/016997, WO 2001/090129, and/or U.S. Pat. No. 6,113,918 and have been reported in the literature. Additional AGP derivatives are disclosed in U.S. Pat. Nos. 7,129,219, 6,525,028 and 6,911,434. Certain AGPs act as agonists of TLR4, while others are recognized as TLR4 antagonist

In addition to the immunostimulatory agents described above, the compositions of the present invention may further comprise other therapeutic agents which, because of their adjuvant nature, can act to stimulate the immune system to respond to the cancer antigens present on the inactivated tumor cell(s). Such adjuvants include, but are not limited to, lipids, liposomes, inactivated bacteria which induce innate immunity (e.g., inactivated or attenuated Listeriamonocytogenes), compositions which mediate innate immune activation via, (NOD)-like receptors (NLRs), Retinoic acid inducible gene-based (RIG)-I-like receptors (RLRs), and/or C-type lectin receptors (CLRs). Examples of PAMPs include lipoproteins, lipopolypeptides, peptidoglycans, zymosan, lipopolysaccharide, neisserial porins, flagellin, profillin, galactoceramide, muramyl dipeptide. Peptidoglycans, lipoproteins, and lipoteichoic acids are cell wall components of Gram-positive bacteria. Lipopolysaccharides are expressed by most bacteria, with MPL being one example. Flagellin refers to the structural component of bacterial flagella that is secreted by pathogenic and commensal bacteria, rt-Galactosylceramide (rt.-GalCer) is an activator of natural killer T (NKT) cells. Muramyl dipeptide is a bioactive peptidoglycan motif common to all bacteria.

Because of their adjuvant qualities, TLR agonists are preferably used in combinations with other vaccines, adjuvants and/or immune modulators, and may be combined in various combinations. Thus, in certain embodiments, the herein described compounds of Formula I that bind to STING and induce STING-dependent TBKI activation and an inactivated tumor cell which expresses and secretes one or more cytokines which stimulate DC induction, recruitment and/or maturation, as described herein can be administered together with one or more TLR agonists for therapeutic purposes.

Indoleamine 2,3-dioxygenase 1 (IDO1) is a key immunosuppressive enzyme that modulates the anti-tumor immune response by promoting regulatory T cell generation and blocking effector T cell activation, thereby facilitating tumor growth by allowing cancer cells to avoid immune surveillance (Lemos 2016; Munn 2016). Further active ingredients (antineoplastic agents) for use in combination or co-administered with the presently invented compounds of Formula I are IDO inhibitors. Epacadostat, ((Z)—N-(3-bromo-4-fluorophenyl)-N′-hydroxy-4-[2-(sulfamoylamino)ethylamino]-1,2,5-oxadiazole-3-carboxamidine) is a highly potent and selective oral inhibitor of the IDO1 enzyme that reverses tumor-associated immune suppression and restores effective anti-tumor immune responses. Epacadostat is disclosed in U.S. Pat. No. 8,034,953.

Additional examples of other therapeutic agents (anti-neoplastic agent) for use in combination or co-administered with a compound of Formula I are CD73 inhibitors and A2a and A2b adenosine antagonists.

The compounds of Formula I and pharmaceutically acceptable salts thereof may be used in combination with at least one other therapeutic agent useful in the prevention or treatment of bacterial and viral infections. Examples of such agents include, without limitation: polymerase inhibitors such as those disclosed in WO 2004/037818, as well as those disclosed in WO 2004/037818 and WO 2006/045613; JTK-003, JTK-019, NM-283, HCV-796, R-803, R1728, R1626, as well as those disclosed in WO 2006/018725, WO 2004/074270, WO 2003/095441, US2005/0176701, WO 2006/020082, WO 2005/080388, WO 2004/064925, WO 2004/065367, WO 2003/007945, WO 02/04425, WO 2005/014543, WO 2003/000254, EP 1065213, WO 01/47883, WO 2002/057287, WO 2002/057245 and similar agents; replication inhibitors such as acyclovir, famciclovir, ganciclovir, cidofovir, lamivudine and similar agents; protease inhibitors such as the HIV protease inhibitors saquinavir, ritonavir, indinavir, nelfinavir, amprenavir, fosamprenavir, brecanavir, atazanavir, tipranavir, palinavir, lasinavir, and the HCV protease inhibitors BILN2061, VX-950, SCH503034; and similar agents; nucleoside and nucleotide reverse transcriptase inhibitors such as zidovudine, didanosine, lamivudine, zalcitabine, abacavir, stavudine, adefovir, adefovir dipivoxil, fozivudine, todoxil, emtricitabine, alovudine, amdoxovir, elvucitabine, tenofovir disproxil fumarate, tenofovir alafenamide fumarate/hemifumarate, and similar agents; non-nucleoside reverse transcriptase inhibitors (including an agent having anti-oxidation activity such as immunocal, oltipraz etc.) such as nevirapine, delavirdine, efavirenz, loviride, immunocal, oltipraz, capravirine, TMC-278, TMC-125, etravirine, rilpivirine and similar agents; entry inhibitors such as enfuvirtide (T-20), T-1249, PRO-542, PRO-140, TNX-355, BMS-806, 5-Helix and similar agents; integrase inhibitors such as dolutegravir, elvitegravir, raltegravir L-870,180 and similar agents; budding inhibitors such as PA-344 and PA-457, and similar agents; chemokine receptor inhibitors such as vicriviroc (Sch-C), Sch-D, TAK779, maraviroc (UK-427,857), TAK449, as well as those disclosed in WO 02/74769, WO 2004/054974, WO 2004/055012, WO 2004/055010, WO 2004/055016, WO 2004/055011, and WO 2004/054581, and similar agents; pharmacokinetic enhancers such as cobicistat; neuraminidase inhibitors such as CS-8958, zanamivir, oseltamivir, peramivir and similar agents; ion channel blockers such as amantadine or rimantadine and similar agents; and interfering RNA and antisense oligonucleotides and such as ISIS-14803 and similar agents; antiviral agents of undetermined mechanism of action, for example those disclosed in WO 2005/105761, WO 2003/085375, WO 2006/122011, ribavirin, and similar agents.

The compounds of Formula I and pharmaceutically acceptable salts thereof may also be used in combination with other therapeutic agents which may be useful in the treatment of Kaposi's sarcoma-associated herpesvirus infections (KSHV and KSHV-related) include, without limitation chemotherapeutic agents such as bleomycin, vinblastine, vincristine, cyclophosphamide, prednisone, alitretinoin and liposomal anthracyclines such as doxorubicin, daunorubicin, immunotherapeutics such as Rituximab, Tocilizumab, Siltuximab and others such as Paclitaxel and Rapamycin.

In one embodiment of this invention, the at least one other therapeutic agent is an antimycobacterial agent or a bactericidal antibiotic. The compounds of Formula I and pharmaceutically acceptable salts thereof may also be used in combination with at least one other therapeutic agent which may be useful in the treatment of TB infection Mycobacterium tuberculosis) and Tularemia (Franciseiia tularensis) include without limitation to first line oral agents isoniazid, Rifampicin, pyrazinamide, ethambutol, streptomycin, rifabutin; injectable agents including kanamycin, amikacin, capreomycin, streptomycin; fluoroquinolones including levofloxacin moxifloxacin ofloxacin; oral bacteriostatic agents para-aminosalicylic acid cycloserine terizidone thionamide protionamide; SQ-109 PNU-100480, Rifapentine Linezolid, PA-824 AZD5847, Gatifloxacin Moxifloxacin, Sirturo (bedaquiline) Delamanid (OPC-67683) and agents with undetermined mechanism of action in the treatment of drug-resistant TB, including clofazimine, linezolid, amoxicillin/clavulanate thioacetazone imipenem/cilastatin high dose isoniazid clarithromycin, ciprofloxacin. The compounds of Formula I and pharmaceutically acceptable salts thereof may also be used in combination with an antimycobacterial agent (such as isoniazid (INH), ehambutol (Myambutol®), rifampin (Rifadin®), and pyrazinamide (PZA)) a bactericidal antibiotic (such as rifabutin (Mycobutin®) or rifapentine (Priftin®)), an aminoglycoside (Capreomycin®), a fluorquinolone (levofloxacin, moxifloxicin, ofloxacin), thioamide (ehionamide), cyclosporine (Sandimmune®), para-aminosalicyclic acid (Paser®), cycloserine (Seromycin®), kanamycin (Kantrex®), streptomycin, viomycin, capreomycin (Capastat®)), bedaquiline fumarate (Sirturo®), oxazolidinone (Sutezolid®), PNU-100480, or delamanid (OPC-67683).

The compounds of Formula I and pharmaceutically acceptable salts thereof may also be used in combination with at least one other therapeutic agent which may be useful in the treatment of Chlamydia include, without limitations Azithromycin, Doxycycline, Erythromycin, Levofloxacin, Ofloxacin.

The compounds of this invention may also be used in combination with at least one other therapeutic agent which may be useful in the treatment of Plasmodium infection include, without limitations to chloroquine, atovaquone-proguanil, artemether-lumefantrine, mefloquine, quinine, quinidine, doxocycline, cindamycin, artesunate, primaquine.

In the treatment of amyotrophic lateral sclerosis (ALS), a compound of Formula I or a pharmaceutically acceptable salts thereof may be used in combination with a glutamate blocker (Riluzole (Rilutek®)), quinidine (Nuedexta 6), anticholinergics (Amitriptyline®, Artane®, scopolamine patch (Transderm Scop®)), sympathomimetics (pseudoephedrine), mucolytics (guaifenesin), or analgesics (tramadol (Ultram®); ketorolac (Toradol®); morphine; fentanyl patch (Duragesic®)).

In the treatment of multiple scelrosis, a compound of Formula I or pharmaceutically acceptable salts thereof may be used in combination with corticosteroids (prednisone, methylprednisolone), Interferon Beta 1-A (Avonex®, Extavia®, Rebif®, Betaseron®), peginterferon beta-IA (Plegridy®), Glatiramer acetate (Copaxone®); glatiramer acetate (Glatopa®-generic equivalent of Copaxone); Dimethyl fumarate (Tecfidera®); Fingolimod (Gilenya®)); teriflunomide (Aubagio®); dalfampridine (Ampyra®); daclizumab (Zinbryta); alemtuzumab (Lemtrada®); natalizumab (Tysabri®); or mitoxantrone hydrochloride (Novantrone®).

The compounds of this invention may also be used as adjuvants to improve the immune response raised to any given antigen and/or reduce reactogenicity/toxicity in a patient, particularly a human, in need thereof. As such, a compound of this invention may be used in combination with vaccine compositions to modify, especially to enhance, the immune response for example by increasing the level or duration of protection and/or allowing a reduction in the antigenic dose.

The compounds of Formula I and pharmaceutically acceptable salts thereof may be used in combination with one or more vaccines or immugenic antigens useful in the prevention or treatment of viral infections. Such vaccines or immugenic antigens include, without limitation to pathogen derived proteins or particles such as attenuated viruses, virus particles, and viral proteins typically used as immugenic substances. Examples of viruses and viral antigens include, without limitations to Polioviruses, Coronaviridae and Coronaviruses, Rhinovirus (all subtypes), Adenoviruses (all subtypes), Hepatitis A, Hepatitis B, Hepatitis C, Hepatitis D, Human papillomavirus (including all subtypes), Rabies viruses, Human T-cell lympotropic virus (all subtypes), Rubella virus, Mumps virus, Coxsackie virus A (all subtypes), Cosackie virus B (all subtypes), human enteroviruses, herpesviruses including cytomegalovirus, Epstein-Barr virus, human herepesvi ruses (all subtypes), herpes simplex virus, varicella zoster virus, human immunodeficiency virus (HIV) (all subtypes), Epstein-Barr virus, Reoviruses (all subtypes), Filoviruses including Marburg virus and Ebola virus (all stains), Arenaviruses including Lymphocytic choriomeningitis virus, Lassa virus, Junin virus, and Machupo virus, Arboviruses including West Nile virus, Dengue viruses (all serotypes), Zika virus, Colorado tick fever virus, Sindbis virus, Togaviraidae, Flaviviridae, Bunyaviridae, Reoviridae, Rhabdoviridae, Orthomyxoviridae, Poxviruses including orthopoxvirus (variola virus, monkypox virus, vaccinia virus, cowpox virus), yatapoxviruses (tanapox virus, Yaba monkey tumor virus), parapoxvirus, molluscipoxvirus, Yellow fever, Hantaviruses including Hantaan, Seoul, Dobrava, Sin Nombre, Puumala, and Dobrava-like Saaremaa, human para influenza viruses and influenza viruses (all types), H1N1 influenza and swine influenza viruses, respiratory syncytial virus (all subgroups), rotaviruses including human rotaviruses A-E, bovine rotavirus, rhesus monkey rotavirus, Polyomaviruses including simian virus 40, JC virus, BK virus, Coltiviruses, eyach virus, calciviruses, and Parvoviridae including dependovirus, parvovirus and erythrovirus.

Accordingly, this invention provides an immugenic composition comprising an antigen or antigenic composition and a compound of Formula I, or a pharmaceutically acceptable salt thereof. There is further provided a vaccine composition comprising an antigen or antigenic composition and a compound of Formula I, or a pharmaceutically acceptable salt thereof. The compounds of Formula I and pharmaceutically acceptable salts thereof may also be used in combination with at least one other therapeutic agent which may be useful in the prevention or treatment of viral infections for example immune therapies (e.g. interferon or other cytokines/chemokines, cytokine/chemokine receptor modulators, cytokine agonists or antagonists and similar agents); and therapeutic vaccines, anti-fibrotic agents, antiinflammatory agents such as corticosteroids or NSAIDs (non-steroidal anti-inflammatory agents) and similar agents.

A compound that modulates STING, particularly a compound of Formula I or a pharmaceutically acceptable salt thereof, may be administered in combination with other antiinflammatory agents, including oral or topical corticosteroids, anti-TNF agents, 5-aminosalicyclic acid and mesalamine preparations, hydroxycloroquine, thiopurines, methotrexate, cyclophosphamide, cyclosporine, calcineurin inhibitors, mycophenolic acid, mTOR inhibitors, JAK inhibitors, Syk inhibitors, anti-inflammatory biologic agents, including anti-IL6 biologies, anti-ILI agents, anti-IL17 biologies, anti-CD22, anti-integrin agents, anti-IFNα, anti-CD20 or CD4 biologies and other cytokine inhibitors or biologies to T-cell or B-cell receptors or interleukins.

For example, in the treatment of systemic lupus erythematosus and related lupus disorders, a compound that modulates STING, particularly a compound of Formula I, or a pharmaceutically acceptable salt thereof, may be administered in combination with at least one other therapeutic agent, including, a corticosteroid (such as prednisolone (Delatsone®, Orapred, Millipred, Omnipred, Econopred, Flo-Pred), an immunosuppressive agent (such as methotrexate (Rhuematrex®, Trexall®), dexamethasone (Decadron®, Solurex®), Mycophenolate mofetil (Cellcept®), Tacrolimus®, Sirolimus®), B-cell therapy (belimumab (Benlysta®), B-cell inhibitor (Atacicept®, Apratuzumab® (anti-CD22), SBI-087 (anti-CD20), an anti-BAFF antibody (LY2127399, A623), Velcade®), azathioprine (Azasan®, Imuran®), triamcinolone (Clinacort®, Kenalog-10®), hydroxychloroquine (Plaquenil®), thalidomide (Immunoprin®, Contergan®), immunoglobulin therapy (HyQiva®, Flebogamma®, Gamunex®, Privigen®, Gammagard®), anti-interferon-alpha therapy (Rontalizumab®, Sifalimumab®, AGS-009®, IFN Kinoid), TLR7 and TLR9 blockers (IMO-3100), anti-cytokine therapies (anti-IL6 (CNTO-136), anti-interferon-gamma (AMG811), immunomodulatory therapy (Lupuzor™, Abatacept, Orencia®, AMG557, Laquinimod, Paquinimod, Leflunomide, anti-ICOS (Medi-570), anti-CD40 ligand antibody (CDP7657)), and/or a platelet aggregation inhibitor (aspirin).

In treatment of vasculitis and disease with inflammation of small or medium size blood vessels, a compound that modulates STING, particularly a compound of Formula I, or a pharmaceutically acceptable salt thereof, may be administered in combination with alkylating agents (cyclophosphamide, Cytoxan®), anti-rheumatic anti-CD20 antibody (Rituxan®, Rituximab®), and anti-TNF inhibitors (Etanrcept®).

In the treatment of psoriasis, a compound that modulates STING, particularly a compound of Formula I, or a pharmaceutically acceptable salt thereof, may be administered in combination with ixekizumab, tildrakizumab (MK-3222), or secukinumab (AIN₄₅₇).

In one embodiment of this invention, the at least one other therapeutic agent is selected from an inhaled corticosteroid, a long acting beta agonist, a combination of an inhaled corticosteroid and a long acting beta agonist, a short acting beta agonist, a leukotriene modifier, an anti-IgE, a methylxanthine bronchodilator, a mast cell inhibitor, and a long-acting muscarinic antagonist. For example, in the treatment of asthma, a compound that inhibits STING, particularly a compound of Formula I or a pharmaceutically acceptable salt thereof, may be administered in combination with an inhaled corticosteroid ((ICS) such as fluticasone proprionate (Flovent®), beclomethasone dipropionate (QVAR®), budesonide (Pulmicort), trimcinolone acetonide (Azmacort®), flunisolide (Aerobid®), mometasone fuorate (Asmanex® Twisthaler®), or Ciclesonide (Alvesco®)), a long acting beta agonist ((LABA) such as formoterol fumarate (Foradil®), salmeterol xinafoate (Serevent®)), a combination of an ICS and LABA (such as fluticasone furoate and vilanterol (Breo Ellipta®), formoterol/budesonide inhalation (Symbicort®), beclomethasone dipropionate/formoterol (Inuvair®), and fluticasone propionate/salmeterol (Advair®), a short acting beta agonist ((SABA) such as albuterol sulfate (ProAir®, Proventil HFA®, Ventolin HFA®, AccuNeb® Inhalation Solution), levalbuterol tartrate (Xopenex® HFA), ipratropium bromide/albuterol (Combivent® Respimat®), ipratropium bromide (Atrovent® HFA), a leukotriene modifier (such as montelukast sodium (Singulair®), zafirlukast (Accolate®), or zileuton (Zyflo®), and anti-IgE (such as omalizumab (Xolair®)), a methylxanthine bronchodilator (such as theophylline (Accurbron®, Aerolate®, Aquaphyllin®, Asbron®, Bronkodyl®, Duraphyl®, Elixicon®, Elixomin®, Elixophyllin®, Labid®, Lanophyllin®, Quibron-T®, Slo-Bid®, Slo-Phyllin®, Somophyllin®, Sustaire®, Synophylate®, T-Phyll®, Theo-24®, Theo-Dur®, Theobid®, Theochron®, Theoclear®, Theolair®, Theolixir®, Theophyl®, Theovent®, Uni-dur®, Uniphyl®), a mast cell inhibitor (such as cromulyn sodium (Nasalcrom®) and nedocromil sodium (Tilade®)), a long-acting muscarinic antagonist ((LAMA) such as mometasone furoate/formoterol fumarate dihydrate (Dulera®)).

Other agents that may be suitable for use in combination therapy in the treatment of asthma include a protein tyrosine kinase inhibitor (masitinib), CRTH2/D-prostanoid receptor antangonist (AMG 853), indacaterol (Arcapta® Neohaler®), an epinephrine inhalation aerosol (E004), fluticasone furoate/fluticasone proprionate, vinanterol inhalation/fluticasone furoate powder (Relovair™), fluticasone propionate/eformoterol fumarate dehydrate (Flutiform®), reslizumab, salbutamol dry-powder inhalation, tiotropium bromide (Spiriva® HandiHaler®), formoterol/budesonide (Symbicort® SMART®), fluticasone furoate (Veramyst®), Vectura's VR506, lebrikizumab (RG3637), a combination phosphodiesterase (PDE)-3 and (PDE)-4 inhibitor (RPL554).

In one embodiment of this invention, the at least one other therapeutic agent is selected from a long acting beta agonist, a long-acting inhaled anticholinergic or muscarinic antagonist, a phosphodiesterase inhibitor, a combination of an inhaled corticosteroid long acting beta agonist, a short acting beta agonist, and an inhaled corticosteroid. For example, in the treatment of COPD, a compound that modulates STING, particularly a compound of Formula I or a pharmaceutically acceptable salt thereof, may be administered in combination with a LABA (such as salmeterol xinafoate (Serevent), umeclidinium/vilanterol (Anoro Ellipta®), umeclidinium (Incruse Ellipta®), aformoterol tartrate (Brovana®), formoterol fumarate inhalation powder (Foradil®), indacterol maleate (Arcapta® Neohaler®), or fluticasone propionate/eformoterol fumarate dehydrate (Flutiform®)), a long-acting inhaled anticholinergic (or muscarinic antagonist, such as tiotropium bromide (Spiriva®), and aclidinium bromide (Tudorza® Pressair®), a phosphodiesterase (PDE-r) inhibitor (such as roflumilast, Daliresp®), a combination ICS/LABA (such as fluticasone furoate and vilanterol (Breo Ellipta®), fluticasone propionate/salmeterol (Advair®), budesonide/formoterol (Symbicort®), mometasone/formoterol (Dulera®), ipratropium bromide/albuterol sulfate (Duoneb®, Atrovent®), albuterol/ipratropium (Combivent Respimat®)), a SABA (such as ipratropium bromide (Atrovent®), and albuterol sulfate (ProAir®, Proventil®)), and an ICS (such as budesonide (Pulmicort®) and fluticasone propionate (Flovent®), beclometasone dipropionate (QVAR®).

Other agents that may be suitable for use in combination therapy in the treatment of COPD include SCH527123 (a CXCR2 antagonist), glycoprronium bromide ((NVA237) Seebri® Breezhaler®), glycopyrronium bromide and indacaterol maleate ((QVA149) Ultibro® Breezhaler®), glycopyrrolate and formoterol fumarate (PT003), indacaterol maleate (QVA149), olodaterol (Striverdi® Respimat®), tiotropium (Spiriva®)/olodaterol (Striverdi® Respimat®), and aclidinium/formoterol inhalation.

In one embodiment of this invention, the at least one other therapeutic agent is selected from an oral corticosteroid, anti-thymocyte globulin, thalidomide, chlorambucil, a calcium channel blocker, a topical emollient, an ACE inhibitor, a serotonin reuptake inhibitor, an endothelin-1 receptor inhibitor, an anti-fibrotic agent, a proton-pump inhibitor or imatinib, ARG201, and tocilizumab. For example, in the treatment of systemic scleroderma, a compound that modulates STING, particularly a compound of Formula I or a pharmaceutically acceptable salt thereof, may be administered in combination with an oral corticosteroid (such as prednisolone (Delatsone®, Orapred, Millipred, Omnipred, Econopred, Flo-Pred), an immunosuppressive agent (such as methotrexate (Rhuematrex®, Trexall®), cyclosporine (Sandimmune®), anti-thymocyte globulin (Atgam®), mycophenolate mofetil (CellCept®), cyclophosphamide (Cytoxan®), FK506 (tacrolimus), thalidomide (Thalomid®), chlorambucil (Leukeran®), azathioprine (Imuran®, Azasan®)), a calcium channel blocker (such as nifedipine (Procardia®, Adalat®) or nicardipine (Cardene®), a topical emollient (nitroglycerin ointment), an ACE inhibitor (such as lisinopril (Zestril®, Prinivil®), diltaizem (Cardizem®, Cardizem SR®, Cardizem CD®, Cardia®, Dilacor®, Tiazac®)), a serotonin reuptake inhibitor (such as fluoxetine (Prozac®)), an endothelin-1 receptor inhibitor (such as bosentan (Tracleer®) or epoprostenol (Flolan®, Veletri®, Prostacyclin®)) an anti-fibrotic agent (such as colchicines (Colcrys®), para-aminobenzoic acid (PABA), dimethyl sulfoxide (DMSO), and D-penicillamine (Cuprimine®, Depen®), interferon alpha and interferon gamma (INF-g)), a proton-pump Inhibitor (such as omeprazole (Prilosec®), metoclopramide (Reglan®), lansoprazole (Prevacid®), esomeprazole (Nexium®), pantoprazole (Protonix®), rabeprazole (Aciphex®)) or imatinib (Gleevec®) ARG201 (arGentis Pharmaceutical), belimumab (Benlysta®), tocilizumab (Actema®).

In the treatment of Sjogren's syndrome, a compound that modulates STING, particularly a compound of Formula I or a pharmaceutically acceptable salt thereof, may be administered in combination with anti-rheumatic agents (hydroxychloroquine and Plaquenil®, Ridaura®, Kineret®), cholinergic agonists (Salagen®, Evoxac®), a JAK inhibitor (Xelijanz®, and anti-TNF treatments (Remicade®, Humira®, Enbrel®, Cimzia®, Simponi®).

In one embodiment of this invention, the at least one other therapeutic agent is a ciliary neurotrophic growth factor or a gene transfer agent. For example, in the treatment of retinitis pigmentosa, a compound that modulates STING, particularly a compound of Formula I or a pharmaceutically acceptable salt thereof, may be administered in combination with a ciliary neurotrophic growth factor (NT-501-CNTF) or gene transfer agent, UshStat®.

In one embodiment of this invention, the at least one other therapeutic agent is selected from a trivalent (I1V3) inactivated influenza vaccine, a quadrivalent (I1V4) inactivated influenza vaccine, a trivalent recombinant influenza vaccine, a quadrivalent live attenuated influenza vaccine, an antiviral agent, or inactivated influenza vaccine. For example, in the treatment of influenza, a compound that modulates STING, particularly a compound of Formula I or a pharmaceutically acceptable salt thereof, may be administered in combination with a trivalent (11V3) inactivated influenza vaccine (such as Afluria®, Fluarix®, Flucelvax®, FluLaval®, Fluvirin®, Fluzone®), a quadrivalent (I1V4) inactivated influenza vaccine (such as Fluarix® Quadrivalent, Flulaval® Quadrivalent, Fluzone® Quadrivalent), a trivalent recombinant influenza vaccine (such as FluBlok®), a quadrivalent live attenuated influenza vaccine (such as FluMist® Quadrivalent), an antiviral agent (such as oseltamivir (Tamiflu®), zanamivir (Relenza®), rimantadine (Flumadine®), or amantadine (Symmetrel®)), or Fluad®, Fludase, FluNhance®, Preflucel, or VaxiGrip®

In the treatment of a staphylococcus infection, a compound that modulates STING, particularly a compound of Formula I or a pharmaceutically acceptable salt thereof, may be administered in combination with an antibiotic (such as a β-Lactam cephalosporin (Duricef®, Kefzol®, Ancef®, Biocef®, etc), nafcillin (Unipen®), a sulfonamide (sulfamethoxazole and trimethoprim (Bacrim®, Septra®,) sulfasalazine (Azulfidine®), acetyl sulfisoxazole (Gantrisin®), etc), or vancomycin (Vancocin®)).

In one embodiment of this invention, the at least one other therapeutic agent is selected from a topical immunomodulator or calcineurin inhibitor, a topical corticosteroid, an oral corticosteroid, an interferon gamma, an antihistamine, or an antibiotic. For example, in the treatment of atopic dermatitis, a compound that modulates STING, particularly a compound of Formula I, or a pharmaceutically acceptable salt thereof, may be administered in combination with a topical immunomodulator or calcineurin inhibitor (such as pimecrolimus (Elidel®) or tacrolimus ointment (Protopic®)), a topical corticosteroid (such as hydrocortizone (Synacort®, Westcort®), betamethasone (Diprolene®), flurandrenolide (Cordan®), fluticasone (Cutivate®), triamcinolone (Kenalog®), fluocinonide (Lidex®), and clobetasol (Temovate®)), an oral corticosteroid (such as hydrocortisone (Cortef®), methyl prednisolone (Medrol®), or prednisolone (Pediapred®, Prelone®), an immunosuppressant (such as cyclosporine (Neoral®) or interferon gamma (Alferon N®, Infergen®, Intron A, Roferon-A®)), an antihistamine (for itching such as Atarax®, Vistaril®, Benadryl®), an antibiotic (such as penicillin derivatives flucloxacillin (Floxapen®) or dicloxacillin (Dynapen®), erythromycin (Eryc®, T-Stat®, Erythra-Derm®, etc.)), a non-steroidal immunosuppressive agent (such as azathioprine (Imuran®, Azasan®), methotrexate (Rhuematrex®, Trexall®), cyclosporin (Sandimmune®), or mycophenolate mofetil (CellCept®)).

The compounds of the invention may also be formulated with vaccines as adjuvants to modulate their activity. Such compositions may contain antibody(ies) or antibody fragment(s) or an antigenic component including but not limited to protein, DNA, live or dead bacteria and/or viruses or virus-like particles, together with one or more components with adjuvant activity including but not limited to aluminum salts, oil and water emulsions, heat shock proteins, lipid A preparations and derivatives, glycolipids, other TLR agonists such as CpG DNA or similar agents, cytokines such as GM-CSF or IL-12 or similar agents.

In a further aspect of the invention, there is provided a vaccine adjuvant comprising a compound of Formula I, or a pharmaceutically acceptable salt thereof. There is further provided a vaccine composition comprising a compound of Formula I, or a pharmaceutically acceptable salt thereof, and an antigen or antigen composition.

Methods of Treatment

The compounds of the present invention may be used in a method of therapy. Also provided is a method of treatment, comprising administering to a subject in need of treatment a therapeutically-effective amount of a compound of the invention. The term “therapeutically effective amount” is an amount sufficient to show benefit to a patient. Such benefit may be at least amelioration of at least one symptom. The actual amount administered, and rate and time-course of administration, will depend on the nature and severity of what is being treated. Prescription of treatment, e.g. decisions on dosage, is within the responsibility of general practitioners and other medical doctors.

Administration

The active compound or pharmaceutical composition comprising the active compound may be administered to a subject by any convenient route of administration, whether systemically/peripherally or at the site of desired action, including but not limited to, oral (e.g. by ingestion); topical (including e.g. transdermal, intranasal, ocular, buccal, and sublingual); pulmonary (e.g. by inhalation or insufflation therapy using, e.g. an aerosol, e.g. through mouth or nose); rectal; vaginal; parenteral, for example, by injection, including subcutaneous, intradermal, intramuscular, intravenous, intraarterial, intracardiac, intrathecal, intraspinal, intracapsular, subcapsular, intraorbital, intraperitoneal, intratracheal, subcuticular, intraarticular, subarachnoid, intravitreal and intrasternal; by implant of a depot, for example, subcutaneously, intravitreal or intramuscularly. The subject may be a eukaryote, an animal, a vertebrate animal, a mammal, a rodent (e.g. a guinea pig, a hamster, a rat, a mouse), murine (e.g. a mouse), canine (e.g. a dog), feline (e.g. a cat), equine (e.g. a horse), a primate, simian (e.g. a monkey or ape), a monkey (e.g. marmoset, baboon), an ape (e.g. gorilla, chimpanzee, orangutan, gibbon), or a human.

In one embodiment where treating tumours, intratumoural injection may used.

Formulations

While it is possible for the active compound or combination of compounds to be administered alone, it is preferable to present it as a pharmaceutical composition (e.g. formulation) comprising at least one active compound, as defined above, together with one or more pharmaceutically acceptable carriers, adjuvants, excipients, diluents, fillers, buffers, stabilisers, preservatives, lubricants, or other materials well known to those skilled in the art and optionally other therapeutic or prophylactic agents.

Thus, the present invention further provides pharmaceutical compositions, as defined above, and methods of making a pharmaceutical composition comprising admixing at least one active compound, as defined above, together with one or more pharmaceutically acceptable carriers, excipients, buffers, adjuvants, stabilisers, or other materials, as described herein.

The term “pharmaceutically acceptable” as used herein pertains to compounds, materials, compositions, and/or dosage forms which are, within the scope of sound medical judgement, suitable for use in contact with the tissues of a subject (e.g. human) without excessive toxicity, irritation, allergic response, or other problem or complication, commensurate with a reasonable benefit/risk ratio. Each carrier, excipient, etc. must also be “acceptable” in the sense of being compatible with the other ingredients of the formulation.

Suitable carriers, excipients, etc. can be found in standard pharmaceutical texts, for example, Remington's Pharmaceutical Sciences, 18th edition, Mack Publishing Company, Easton, Pa., 1990.

The formulations may conveniently be presented in unit dosage form and may be prepared by any methods well known in the art of pharmacy. Such methods include the step of bringing into association the active compound with the carrier which constitutes one or more accessory ingredients. In general, the formulations are prepared by uniformly and intimately bringing into association the active compound with liquid carriers or finely divided solid carriers or both, and then if necessary shaping the product.

Formulations may be in the form of liquids, solutions, suspensions, emulsions, elixirs, syrups, tablets, losenges, granules, powders, capsules, cachets, pills, ampoules, suppositories, pessaries, ointments, gels, pastes, creams, sprays, mists, foams, lotions, oils, boluses, electuaries, or aerosols.

Formulations suitable for oral administration (e.g. by ingestion) may be presented as discrete units such as capsules, cachets or tablets, each containing a predetermined amount of the active compound; as a powder or granules; as a solution or suspension in an aqueous or non-aqueous liquid; or as an oil-in-water liquid emulsion or a water-in-oil liquid emulsion; as a bolus; as an electuary; or as a paste.

A tablet may be made by conventional means, e.g., compression or moulding, optionally with one or more accessory ingredients. Compressed tablets may be prepared by compressing in a suitable machine the active compound in a free-flowing form such as a powder or granules, optionally mixed with one or more binders (e.g. povidone, gelatin, acacia, sorbitol, tragacanth, hydroxypropylmethyl cellulose); fillers or diluents (e.g. lactose, microcrystalline cellulose, calcium hydrogen phosphate); lubricants (e.g. magnesium stearate, talc, silica); disintegrants (e.g. sodium starch glycolate, cross-linked povidone, cross-linked sodium carboxymethyl cellulose); surface-active or dispersing or wetting agents (e.g. sodium lauryl sulfate); and preservatives (e.g. methyl p-hydroxybenzoate, propyl p-hydroxybenzoate, sorbic acid). Moulded tablets may be made by moulding in a suitable machine a mixture of the powdered compound moistened with an inert liquid diluent. The tablets may optionally be coated or scored and may be formulated so as to provide slow or controlled release of the active compound therein using, for example, hydroxypropylmethyl cellulose in varying proportions to provide the desired release profile. Tablets may optionally be provided with an enteric coating, to provide release in parts of the gut other than the stomach.

Formulations suitable for topical administration (e.g. transdermal, intranasal, ocular, buccal, and sublingual) may be formulated as an ointment, cream, suspension, lotion, powder, solution, past, gel, spray, aerosol, or oil. Alternatively, a formulation may comprise a patch or a dressing such as a bandage or adhesive plaster impregnated with active compounds and optionally one or more excipients or diluents.

Formulations suitable for topical administration in the mouth include losenges comprising the active compound in a flavoured basis, usually sucrose and acacia or tragacanth; pastilles comprising the active compound in an inert basis such as gelatin and glycerin, or sucrose and acacia; and mouthwashes comprising the active compound in a suitable liquid carrier.

Formulations suitable for topical administration to the eye also include eye drops wherein the active compound is dissolved or suspended in a suitable carrier, especially an aqueous solvent for the active compound.

Formulations suitable for nasal administration, wherein the carrier is a solid, include a coarse powder having a particle size, for example, in the range of about 20 to about 500 microns which is administered in the manner in which snuff is taken, i.e. by rapid inhalation through the nasal passage from a container of the powder held close up to the nose. Suitable formulations wherein the carrier is a liquid for administration as, for example, nasal spray, nasal drops, or by aerosol administration by nebuliser, include aqueous or oily solutions of the active compound.

Formulations suitable for administration by inhalation include those presented as an aerosol spray from a pressurised pack, with the use of a suitable propellant, such as dichlorodifluoromethane, trichlorofluoromethane, dichloro-tetrafluoroethane, carbon dioxide, or other suitable gases.

Formulations suitable for topical administration via the skin include ointments, creams, and emulsions. When formulated in an ointment, the active compound may optionally be employed with either a paraffinic or a water-miscible ointment base. Alternatively, the active compounds may be formulated in a cream with an oil-in-water cream base. If desired, the aqueous phase of the cream base may include, for example, at least about 30% w/w of a polyhydric alcohol, i.e., an alcohol having two or more hydroxyl groups such as propylene glycol, butane-1,3-diol, mannitol, sorbitol, glycerol and polyethylene glycol and mixtures thereof. The topical formulations may desirably include a compound which enhances absorption or penetration of the active compound through the skin or other affected areas. Examples of such dermal penetration enhancers include dimethylsulfoxide and related analogues.

When formulated as a topical emulsion, the oily phase may optionally comprise merely an emulsifier (otherwise known as an emulgent), or it may comprises a mixture of at least one emulsifier with a fat or an oil or with both a fat and an oil. Preferably, a hydrophilic emulsifier is included together with a lipophilic emulsifier which acts as a stabiliser. It is also preferred to include both an oil and a fat. Together, the emulsifier(s) with or without stabiliser(s) make up the so-called emulsifying wax, and the wax together with the oil and/or fat make up the so-called emulsifying ointment base which forms the oily dispersed phase of the cream formulations.

Suitable emulgents and emulsion stabilisers include Tween 60, Span 80, cetostearyl alcohol, myristyl alcohol, glyceryl monostearate and sodium lauryl sulphate. The choice of suitable oils or fats for the formulation is based on achieving the desired cosmetic properties, since the solubility of the active compound in most oils likely to be used in pharmaceutical emulsion formulations may be very low. Thus the cream should preferably be a non-greasy, non-staining and washable product with suitable consistency to avoid leakage from tubes or other containers. Straight or branched chain, mono- or dibasic alkyl esters such as di-isoadipate, isocetyl stearate, propylene glycol diester of coconut fatty acids, isopropyl myristate, decyl oleate, isopropyl palmitate, butyl stearate, 2-ethylhexyl palmitate or a blend of branched chain esters known as Crodamol CAP may be used, the last three being preferred esters. These may be used alone or in combination depending on the properties required.

Alternatively, high melting point lipids such as white soft paraffin and/or liquid paraffin or other mineral oils can be used.

Formulations suitable for rectal administration may be presented as a suppository with a suitable base comprising, for example, cocoa butter or a salicylate.

Formulations suitable for vaginal administration may be presented as pessaries, tampons, creams, gels, pastes, foams or spray formulations containing in addition to the active compound, such carriers as are known in the art to be appropriate.

Formulations suitable for parenteral administration (e.g. by injection, including cutaneous, subcutaneous, intramuscular, intravenous and intradermal), include aqueous and non-aqueous isotonic, pyrogen-free, sterile injection solutions which may contain anti-oxidants, buffers, preservatives, stabilisers, bacteriostats, and solutes which render the formulation isotonic with the blood of the intended recipient; and aqueous and non-aqueous sterile suspensions which may include suspending agents and thickening agents, and liposomes or exosomes or other microparticulate systems which are designed to target the compound to blood components or one or more organs. Examples of suitable isotonic vehicles for use in such formulations include Sodium Chloride Injection, Ringer's Solution, or Lactated Ringer's Injection. Typically, the concentration of the active compound in the solution is from about 1 ng/mL to about 10 μg/mL, for example from about 10 ng/ml to about 1 μg/mL. The formulations may be presented in unit-dose or multi-dose sealed containers, for example, ampoules and vials, and may be stored in a freeze-dried (lyophilised) condition requiring only the addition of the sterile liquid carrier, for example water for injections, immediately prior to use. Extemporaneous injection solutions and suspensions may be prepared from sterile powders, granules, and tablets. Formulations may be in the form of liposomes or exosomes or other microparticulate systems which are designed to target the active compound to blood components or one or more organs.

Dosage

It will be appreciated by one of skill in the art that appropriate dosages of the compound, and compositions comprising the compound, can vary from patient to patient. Determining the optimal dosage will generally involve the balancing of the level of therapeutic benefit against any risk or deleterious side effects. The selected dosage level will depend on a variety of factors including, but not limited to, the activity of the particular compound, the route of administration, the time of administration, the rate of excretion of the compound, the duration of the treatment, other drugs, compounds, and/or materials used in combination, the severity of the condition, and the species, sex, age, weight, condition, general health, and prior medical history of the patient. The amount of compound and route of administration will ultimately be at the discretion of the physician, veterinarian, or clinician, although generally the dosage will be selected to achieve local concentrations at the site of action which achieve the desired effect without causing substantial harmful or deleterious side-effects.

Administration can be effected in one dose, continuously or intermittently (e.g., in divided doses at appropriate intervals) throughout the course of treatment. Methods of determining the most effective means and dosage of administration are well known to those of skill in the art and will vary with the formulation used for therapy, the purpose of the therapy, the target cell(s) being treated, and the subject being treated. Single or multiple administrations can be carried out with the dose level and pattern being selected by the treating physician, veterinarian, or clinician.

In general, a suitable dose of the active compound is in the range of about 100 ng to about 25 mg (more typically about 1 μg to about 10 mg) per kilogram body weight of the subject per day. Where the active compound is a salt, an ester, an amide, a prodrug, or the like, the amount administered is calculated on the basis of the parent compound and so the actual weight to be used is increased proportionately.

In one embodiment, the active compound is administered to a human patient according to the following dosage regime: about 100 mg, 3 times daily.

In one embodiment, the active compound is administered to a human patient according to the following dosage regime: about 150 mg, 2 times daily.

In one embodiment, the active compound is administered to a human patient according to the following dosage regime: about 200 mg, 2 times daily.

However in one embodiment, the active compound is administered to a human patient according to the following dosage regime: about 50 or about 75 mg, 3 or 4 times daily.

In one embodiment, the active compound is administered to a human patient according to the following dosage regime: about 100 or about 125 mg, 2 times daily.

Treatment

The term “treatment,” as used herein in the context of treating a condition, pertains generally to treatment and therapy, whether of a human or an animal (e.g., in veterinary applications), in which some desired therapeutic effect is achieved, for example, the inhibition of the progress of the condition, and includes a reduction in the rate of progress, a halt in the rate of progress, regression of the condition, amelioration of the condition, and cure of the condition. Treatment as a prophylactic measure (i.e., prophylaxis, prevention) is also included.

The term “therapeutically-effective amount,” as used herein, pertains to that amount of an active compound, or a material, composition or dosage from comprising an active compound, which is effective for producing some desired therapeutic effect, commensurate with a reasonable benefit/risk ratio, when administered in accordance with a desired treatment regimen.

Similarly, the term “prophylactically-effective amount,” as used herein, pertains to that amount of an active compound, or a material, composition or dosage from comprising an active compound, which is effective for producing some desired prophylactic effect, commensurate with a reasonable benefit/risk ratio, when administered in accordance with a desired treatment regimen.

The Subject/Patient

The subject/patient may be an animal, mammal, a placental mammal, a marsupial (e.g., kangaroo, wombat), a monotreme (e.g., duckbilled platypus), a rodent (e.g., a guinea pig, a hamster, a rat, a mouse), murine (e.g., a mouse), a lagomorph (e.g., a rabbit), avian (e.g., a bird), canine (e.g., a dog), feline (e.g., a cat), equine (e.g., a horse), porcine (e.g., a pig), ovine (e.g., a sheep), bovine (e.g., a cow), a primate, simian (e.g., a monkey or ape), a monkey (e.g., marmoset, baboon), an ape (e.g., gorilla, chimpanzee, orangutan, gibbon), or a human.

Furthermore, the subject/patient may be any of its forms of development, for example, a foetus. In one preferred embodiment, the subject/patient is a human.

General Synthesis Methods

The compounds of the invention can be prepared employing the following general methods and using procedures described in detail in the examples. The reaction conditions referred to are illustrative and non-limiting, for example one skilled in the art may use a diverse range of synthetic methods to synthesise the desired compounds such as but not limited to methods described in literature (for example but not limited to March's Advanced Organic Chemistry: Reactions, Mechanisms, and Structure, 7th Edition or Larock's Comprehensive Organic Transformations: Comprehensive Organic Transformations: A Guide to Functional Group Preparations).

Compounds of formula I as described above, can be prepared by synthetic strategies outlined below, wherein the definitions above apply. Thus, in any of the following general synthesis schemes, A¹, A², A³, A⁴, A¹¹, A¹², A¹³, A¹⁴, R¹, R¹¹, R^(C1), R^(C2), R^(C3), R^(C4), R^(C13), R^(C14), R^(P) and Y are as defined for any compound of the invention described herein, including Formula (I) and any embodiment thereof.

General Synthesis 1

Scheme 1a illustrates the synthesis of compounds with the structure G4. One equivalent of aldehyde G1 with substituents R¹, A¹, A², A³ and A⁴ can be reacted with one equivalent of aldehyde G2 with substituents R¹¹, A¹¹, A¹², A¹³ and A¹⁴ and one equivalent of tetra-aniline G3 in the presence of either (a) a suitable acid such as, but not limited to, acetic acid or (b) sodium bisulfite or (c) sodium metabisulfite or (d) Oxone® to give compounds of the type G4. In some instances, two equivalents of the same aldehyde (i.e. G1=G2, R1=R¹¹, A¹=A¹¹, A²=A¹², A³=A¹³, A⁴=A¹⁴)) is reacted with one equivalent of tetra-aniline G3 to give compounds of the type G4. The tetra-aniline may also be symmetrical (R^(C1)=R^(C11), R^(C3)=R^(C13), R^(C4)=R^(C14)). In other instances, the aldehydes G1 and G2 can be reacted with an equivalent of di-nitro/di-aniline compound G5 using conditions such as, but not limited to, heating with sodium dithionite as shown in Scheme 1b.

It will be appreciated by those skilled in the art that as illustrated in schemes 1a and 1b, substituents R^(A) (when A1=CR^(A)), R^(B) (when A²=CR^(B)), R^(C) (when A³=CR^(C)), and R^(D) (when A⁴=CR^(D)), R^(AA) (when A¹¹=CR^(AA)), R^(BB) (when A¹²=CR^(BB)), R^(CC) (when A¹³=CR^(CC)) and R^(DD) (when A¹⁴=CR^(DD)), R^(C1), R^(C3) and R^(C4), R^(C11), R^(C13) and R^(C14) on compounds G1 G5 may be a halogen atom or other suitable leaving group (such as a boronic acid or ester, —OH, —O-triflyl, —OC(O)allyl, and the like) to allow for transition-metal catalysed transformations such as Ulmann, Negishi, Stille, Suzuki-Miyaura, Sonogashira and Buchwald-Hartwig couplings, or for SnAr displacements to produce further examples of the type of compound I.

General Synthesis 2

Compounds of the structure G1 can be accessed by treatment of a suitably substituted arene or heteroarene G6 with either an alkyl lithium such as, but not limited to, n-butyllithium, or a lithium amide base such as, but not limited to, lithium diisopropylamide, and subsequent quenching of the resultant lithiated species with N,N-dimethylformamide. Compounds of the structure G1 may also be accessed via bromination of intermediate G7 to form alkyl bromide G8 using a suitable bromine source such as, but not limited to, N-bromosuccinimide, in the presence of an initiator such as, but not limited to, benzoyl peroxide. An oxidation reaction of G8 may be carried out with a suitable oxidant such as, but no limited to, N-methylmorpholine N-oxide to give compounds of the type G1. Aldehydes of the type G1 can also be prepared via a number of other methods well known to those skilled in the art such as, but not limited to, oxidations of benzyl alcohols, reactions with organolithium compounds, and reductions of acids/esters/nitriles.

General Synthesis 3

Scheme 3 illustrates methods by which compounds of the formula G3 can be made, in which ‘X’ is an appropriate leaving group such as, but not limited to, a halogen atom or activated leaving group such as, but not limited to, triflate, mesylate or tosylate. In Scheme 3, ‘W’ is defined as a primary amide C(═O)NH₂ or a group that can be converted into a primary amide using conditions known by those skilled in the art such as, but not limited to an ester —C(═O)OR^(P), carboxylic acid —CO₂H, activated acid (such as acid halide or mixed anhydride) or nitrile —CN.

The first step in obtaining compounds of the formula G3 is reaction of a phenyl compound substituted with an appropriate leaving group G9 with a di-amine of the type G10 and an equivalent of another phenyl compound substituted with an appropriate leaving group G11 to give the bis-nitro species G12. In some instances, the bis-nitro compound G12 will be symmetrical where R^(C1)=R^(C11), R^(C3)=R^(C13), R^(C4)=R^(C14). Alternatively, compounds of the type G12 can be accessed by first reacting a phenyl compound substituted with an appropriate leaving group G9 with a diamine G13 mono-protected with a suitable protecting group such as, but not limited to, a Boc group to give intermediate G14. Deprotection of the amine using, for example, acidic conditions gives intermediates of the type G15 which can be subsequently reacted with a phenyl compound substituted with an appropriate leaving group G11 to give di-nitro intermediate G12. Reduction of the aromatic nitro groups in compound G12 can be accomplished by methods well known to those skilled in the art, including, but not limited to, catalytic hydrogenation using 10% Pd/C or Raney nickel in a hydrogen atmosphere, iron or zinc in acidic media, or sodium dithionite to give access to tetra-aniline compounds of the type G3.

It will be appreciated by those skilled in the art that substituents R^(C1), R^(C3) and R^(C4), R^(C11), R^(C13) and R^(C14) on compounds G3, G9, G11, G12, G143 and G15 may be a halogen atom or other suitable leaving group (such as a boronic acid or ester, —OH, —O-triflyl, —OC(O)allyl, and the like) to allow for transition-metal catalysed transformations such as Ulmann, Negishi, Stille, Suzuki-Miyaura, Sonogashira and Buchwald-Hartwig couplings, or for SnAr displacements to produce further examples of the type of compound I.

General Synthesis 4

Conditions for conversion of diacid G16 to diester G18 will be apparent to those skilled in the art, but include an excess of a suitable nucleophile G17 and a catalyst such as concentrated sulphuric acid (for formation of an ester), as shown in Scheme 4a. Alternatively G16 may be first activated by a coupling agent such as, but not limited to, 1-[bis(dimethylamino)methylene]-1H-1,2,3-triazolo[4,5-b]pyridinium 3-oxide hexafluorophosphate, 1-ethyl-3-(3-dimethylaminopropyl)carbodiimide or N,N′-dicyclohexylcarbodiimide before reaction with nucleophile G17 in the presence of a catalyst such as, but not limited to, 4-dimethylaminopyridine. Diamide analogues of diester G16 may be prepared by conversion of diacid G16 by conditions apparent to those skilled in the art, including similar conditions as described in Scheme 4a, with a suitably functionalised nucleophile in place of nucleophile G17.

Diester G18 can also be accessed by reaction of diacid G16 with a suitable alkyl halide G19 in the presence of a suitable base such as, but not limited to, cesium carbonate as shown in Scheme 4b.

General Synthesis 5

Scheme 5 illustrates an alternative route into the synthesis of compounds with the structure G4, in which ‘W’ is defined as a primary amide C(═O)NH₂ or a group that can be converted into a primary amide using conditions known by those skilled in the art such as, but not limited to an ester —C(═O)OR^(P), carboxylic acid —CO₂H, activated acid (such as acid halide or mixed anhydride) or nitrile —CN. Nitro intermediate G14 can be reduced to the di-aniline intermediate using conditions such as, but not limited to, stirring over a heterogenous catalyst such as Pd/C under an atmosphere of hydrogen to give G190. Condensation of this di-aniline with aldehyde intermediate G1 in the presence of either (a) a suitable acid such as, but not limited to, acetic acid or (b) sodium bisulfite or (c) sodium metabisulfite or (d) Oxone® provides access to intermediates of the type G20. Treatment of the mono-Boc protected material under acidic conditions liberates the primary amine to give compounds of the type G21. A second nucleophilic aromatic substitution can be performed with a phenyl compound substituted with an appropriate leaving group of the type G11 in the presence of a base such as but not limited to triethylamine to give G22. A reduction step of the nitro compound G22 can be performed to give G23 using conditions such as, but not limited to, stirring over a heterogenous catalyst such as Pd/C under an atmosphere of hydrogen. The di-aniline intermediate G23 can be condensed with an equivalent of an aldehyde of the type G2 in the presence of either (a) a suitable acid such as, but not limited to, acetic acid or (b) sodium bisulfite or (c) sodium metabisulfite or (d) Oxone® to give bis-benzimidazole compounds of the type G4.

General Synthesis 6

Compounds of the formula G26 or G27 can be accessed via the common nitrile intermediate G24 (Scheme 6a). Compounds of the type G24 are accessible using the chemistry described in General Synthesis 1 and General Synthesis 5 where R1/R11 is replaced by a nitrile —CN group. Treatment of a suitably substituted nitrile G24 with hydroxylamine hydrochloride enables access to N-hydroxyamidine intermediates of the type G25. Subsequent treatment of intermediate G25 with a reagent providing a carbonyl (C═O) equivalent such as, but not limited to, carbonyldiimidazole gives 5-oxo-4H-1,2,4-oxadiazol-3-yl derivatives of the type G26. Alternatively, nitrile G24 can be employed to form tetrazole compounds G27 in one step in the presence of a base such as, but not limited to, triethylamine and an azide source such as sodium azide.

Compounds of the type G30 are regioisomers of G26, and can be prepared from acid G16. Coupling of acid G16 with hydrazine substituted on one nitrogen atom with a suitable protecting group (PG′) such as, but not limited to, tert-butyloxycarbonyl (Boc), using coupling agents such as, but not limited to, 1-ethyl-3-(3-dimethylaminopropyl)carbodiimide and hydroxybenzotriazole in the presence of a base such as, but not limited to, N,N-diisopropylethylamine provides access to protected intermediates of the type G28. Removal of the protecting group using conditions well known to those skilled in the art afford compounds of the type G29 which can be reacted with a reagent providing a carbonyl (C═O) equivalent such as, but not limited to, carbonyldiimidazole to give 2-oxo-3H-1,3,4-oxadiazol-5-yl derivatives of the type G30.

Further Embodiments

In some embodiments, the two moeities attached to Y are the same, i.e. A¹¹=A¹, A¹²=A², A¹³=A³, A¹⁴=A⁴, R^(C11)=R^(C1), R^(C13)=R^(C3), R^(C14)=R^(C4), R¹¹=R¹.

A¹-A⁴ and A¹¹-A¹⁴

In some embodiments, A¹ is CR^(A).

In other embodiments, A¹ is N.

In some embodiments, A² is CR^(B).

In other embodiments, A² is N.

In some embodiments, A³ is CR^(C).

In other embodiments, A³ is N.

In some embodiments, A⁴ is CR^(D).

In other embodiments, A⁴ is N.

In some embodiments, two of A¹, A², A³ and A⁴ are N.

In other embodiments, one of A¹, A², A³ and A⁴ are N.

In other embodiments, none of A¹, A², A³ and A⁴ are N, i.e. A¹, A², A³ and A⁴ are CR^(A), CR^(B), CR^(C), and CR^(D) respectively.

In some embodiments, A¹¹ is CR^(AA).

In other embodiments, A¹¹ is N.

In some embodiments, A¹² is CR^(BB).

In other embodiments, A¹² is N.

In some embodiments, A¹³ is CR^(CC).

In other embodiments, A¹³ is N.

In some embodiments, A¹⁴ is CR^(DD).

In other embodiments, A¹⁴ is N.

In some embodiments, two of A¹¹, A¹², A¹³ and A¹⁴ are N.

In other embodiments, one of A¹¹, A¹², A¹³ and A¹⁴ are N.

In other embodiments, none of A¹¹, A¹², A¹³ and A¹⁴ are N, i.e. A¹¹, A¹², A¹³ and A¹⁴ are CR^(AA), CR^(BB), CR^(CC), and CR^(DD) respectively.

In some embodiments, the compound of formula I is selected from compounds of formulae (III)-(IIIe):

R^(A), R^(B), R^(C), R^(D), R^(AA), R^(BB), R^(CC) and R^(DD) (if Present)

In some embodiments, R^(A), R^(B), R^(C), R^(D), R^(AA), R^(BB), R^(CC) and R^(DD), (if present) are selected from H, F, Cl, Br, I, Me, Et, CF₃, cyclopropyl, cyano, OMe, OEt, CH₂OH, CH₂OMe and OH. In some embodiments, R^(A), R^(B), R^(C), R^(D), R^(AA), R^(BB), R^(CC) and R^(DD) (if present) are independently selected from H, F, Cl and Br, preferably H, Cl and F.

In some embodiments one of R^(A), R^(B), R^(C) and R^(D), (if present), and one of R^(AA), R^(BB), R^(CC) and R^(DD), (if present) is selected from H, F, Cl, Br, I, Me, Et, CF₃, cyclopropyl, cyano, OMe, OEt, CH₂OH, CH₂OMe and OH. The remainder (if present) are H.

In other embodiments two of R^(A), R^(B), R^(C) and R^(D), (if present), and two of R^(A), R^(B), R^(C), R^(D), R^(AA), R^(BB), R^(CC) and R^(DD), (if present) are selected from H, F, Cl, Br, I, Me, Et, CF₃, cyclopropyl, cyano, OMe, OEt, CH₂OH, CH₂OMe and OH. The remainder (if present) are H.

In further embodiments three of R^(A), R^(B), R^(C) and R^(D), (if present), and three of R^(A), R^(B), R^(C), R^(D), R^(AA), R^(BB), R^(CC) and R^(DD), (if present) are selected from H, F, Cl, Br, I, Me, Et, CF₃, cyclopropyl, cyano, OMe, OEt, CH₂OH, CH₂OMe and OH. The remainder (if present) are H.

In some embodiments, one, two or three of R^(A), R^(B), R^(C) and R^(D), (if present), and one, two or three of R^(AA), R^(BB), R^(CC) and R^(DD), (if present) are selected from H, F, Cl, Br, Me, CF₃, cyclopropyl, cyano and OMe. The remainder (if present) are H. In some of these embodiments, one, two or three of R^(A), R^(B), R^(C) and R^(D) (if present), and one, two or three of R^(AA), R^(BB), R^(CC) and R^(DD), (if present) are selected from H, F, Cl, Br, Me, CF₃, and cyclopropyl and OMe. The remainder (if present) are H. In some of these embodiments, one, two or three of R^(A), R^(B), R^(C) and R^(D), (if present), and one, two or three of R^(AA), R^(BB), R^(CC) and R^(DD), (if present) are selected from H, F, Cl, Br, Me, and CF₃ and OMe.

In some embodiments one of R^(A), R^(B), R^(C) and R^(D), (if present), and one, two or three of R^(AA), R^(BB), R^(CC) and R^(DD), (if present) is selected from F, Cl, Br, I, Me, Et, CF₃, cyclopropyl, cyano, OMe, OEt, CH₂OH, CH₂OMe and OH. The remainder (if present) are H.

In other embodiments two of R^(A), R^(B), R^(C) and R^(D), (if present), and one, two or three of R^(A), R^(B), R^(C), R^(D), R^(AA), R^(BB), R^(CC) and R^(DD), (if present) are selected from F, Cl, Br, I, Me, Et, CF₃, cyclopropyl, cyano, OMe, OEt, CH₂OH, CH₂OMe and OH. The remainder (if present) are H.

In further embodiments three of R^(A), R^(B), R^(C) and R^(D), (if present), and one, two or three of R^(A), R^(B), R^(C), R^(D), R^(AA), R^(BB), R^(CC) and R^(DD), (if present) are selected from F, Cl, Br, I, Me, Et, CF₃, cyclopropyl, cyano, OMe, OEt, CH₂OH, CH₂OMe and OH. The remainder (if present) are H.

In some embodiments one, two or three of R^(A), R^(B), R^(C) and R^(D), (if present), and one of R^(AA), R^(BB), R^(CC) and R^(DD), (if present) is selected from F, Cl, Br, I, Me, Et, CF₃, cyclopropyl, cyano, OMe, OEt, CH₂OH, CH₂OMe and OH. The remainder (if present) are H.

In other embodiments one, two or three of R^(A), R^(B), R^(C) and R^(D), (if present), and two of R^(A), R^(B), R^(C), R^(D), R^(AA), R^(BB), R^(CC) and R^(DD), (if present) are selected from F, Cl, Br, I, Me, Et, CF₃, cyclopropyl, cyano, OMe, OEt, CH₂OH, CH₂OMe and OH. The remainder (if present) are H.

In further embodiments one, two or three of R^(A), R^(B), R^(C) and R^(D), (if present), and three of R^(A), R^(B), R^(C), R^(D), R^(AA), R^(BB), R^(CC) and R^(DD), (if present) are selected from F, Cl, Br, I, Me, Et, CF₃, cyclopropyl, cyano, OMe, OEt, CH₂OH, CH₂OMe and OH. The remainder (if present) are H.

In some embodiments R^(A) and R^(AA) are selected from H, Br and F, preferably H and F. In some embodiments, R^(A) is H. In some embodiments, R^(AA) is H. In some embodiments R^(A) and

R^(AA) are H. In some embodiments, R^(A) is F. In some embodiments, R^(AA) is F. In some embodiments R^(A) and R^(AA) are F.

In some embodiments R^(B) is selected from H, F, Cl, Br, Me, CF₃, cyclopropyl, cyano, OMe, CH₂OH, and CH₂OMe. In some embodiments R^(BB) is selected from H, F, Cl, Br, Me, CF₃, cyclopropyl, cyano, OMe, CH₂OH, and CH₂OMe. In some embodiments R^(B) and R^(BB) are selected from H, F, Cl, Br, Me, CF₃, cyclopropyl, cyano, OMe, CH₂OH, and CH₂OMe. In some embodiments R^(B) and/or R^(BB) are selected from H, F, Cl, Br, Me, CF₃, cyclopropyl, cyano and OMe. In some embodiments R^(B) and/or R^(BB) are selected from H, F, Br, and C₁. In some embodiments R^(B) and/or R^(BB) are selected from H, F, and Cl. In some embodiments R^(B) and/or R^(BB) are selected from H and C₁. In some embodiments R^(B) and/or R^(BB) are H. In some embodiments R^(B) and/or R^(BB) are F. In some embodiments R^(B) and/or R^(BB) are Cl.

In some embodiments R^(C) is selected from H, F, Cl, Br, Me, CF₃, cyclopropyl, cyano, OMe, CH₂OH, and CH₂OMe. In some embodiments R^(CC) is selected from H, F, Cl, Br, Me, CF₃, cyclopropyl, cyano, OMe, CH₂OH, and CH₂OMe. In some embodiments R^(C) and R^(CC) are selected from H, F, Cl, Br, Me, CF₃, cyclopropyl, cyano, OMe, CH₂OH, and CH₂OMe. In some embodiments R^(C) and/or R^(CC) are selected from H, F, Cl, Br, Me, CF₃, cyclopropyl, cyano and OMe. In some embodiments R^(C) and/or R^(CC) are selected from H, F, Br, and C₁. In some embodiments R^(C) and/or R^(CC) are selected from H, F and C₁. In some embodiments R^(C) and/or R^(CC) are H. In some embodiments R^(C) and/or R^(CC) are F. In some embodiments R^(C) and/or R^(CC) are Cl.

In some embodiments R^(D) is selected from H, F, Cl, Br, Me, CF₃, cyclopropyl, cyano, OMe, CH₂OH, and CH₂OMe. In some embodiments R^(DD) is selected from H, F, Cl, Br, Me, CF₃, cyclopropyl, cyano, OMe, CH₂OH, and CH₂OMe. In some embodiments R^(D) and R^(DD) are selected from H, F, Cl, Br, Me, CF₃, cyclopropyl, cyano, OMe, CH₂OH, and CH₂OMe. In some embodiments R^(D) and/or R^(DD) are selected from H, F, Cl, Br, Me, CF₃, cyclopropyl, cyano and OMe. In some embodiments R^(D) and/or R^(DD) are selected from H, F, Br and C₁. In some embodiments R^(D) and/or R^(DD) are selected from H, F and C₁. In some embodiments R^(D) and/or R^(DD) are H. In some embodiments R^(D) and/or R^(DD) are F. In some embodiments R^(D) and/or R^(DD) are Cl.

In some embodiments, 1 or 2 of R^(A), R^(B), R^(C) and R^(D) (if present) are halo and the remainder are H. In these embodiments, preferred halo atoms are Cl, Br and F, more preferably Cl and F, most preferably F. In these embodiments, 1 or 2 of R^(A) and R^(C) are halo and R^(B) and R^(D) are H.

In some embodiments, 1 or 2 of R^(AA), R^(BB), R^(CC) and A^(DD) (if present) are halo and the remainder are H. In these embodiments, preferred halo atoms are Cl, Br and F, more preferably Cl and F, most preferably F. In these embodiments, 1 or 2 of R^(AA) and R^(CC) are halo and R^(B) and R^(D) are H.

In some embodiments, A¹, A², A³ and A⁴ are selected from combinations 1.1-1.30 in the following table:

Combination A¹ A² A³ A⁴ 1.1 CF CH CH CH 1.2 CH CBr CH CH 1.3 CH CCl CH CH 1.4 CF CCl CH CH 1.5 CH CH CH CBr 1.6 CH CH CCl CH 1.7 CH CF CF CH 1.8 CH CH CH CH 1.9 CF CH CF CH 1.10 CF CH CCl CH 1.11 CBr CH CH CH 1.12 CCl CH CH CH 1.13 N CH CCl CH 1.14 CH CCl CCl CH 1.15 CH CF CH CH 1.16 CH CMe CH CH 1.17 CH N CH CH 1.18 CH CH CBr CH 1.19 CH CH CCF₃ CH 1.20 CH CH CEt CH 1.21 CH CH CF CH 1.22 CH CH CMe CH 1.23 CH CH C—OMe CH 1.24 CH CH CCl CCl 1.25 CH CH CH CF 1.26 CH CH CH C—OMe 1.27 CH CH CH N 1.28 Cl CH CH CH 1.29 CH CH CCN CH 1.30 CBr CH CCl CH

In some embodiments, A¹¹, A¹², A¹³ and A¹⁴ are selected from combinations 2.1-2.30 in the following table:

Combination A¹¹ A¹² A¹³ A¹⁴ 2.1 CF CH CH CH 2.2 CH CBr CH CH 2.3 CH CCl CH CH 2.4 CF CCl CH CH 2.5 CH CH CH CBr 2.6 CH CH CCl CH 2.7 CH CF CF CH 2.8 CH CH CH CH 2.9 CF CH CF CH 2.10 CF CH CCl CH 2.11 CBr CH CH CH 2.12 CCl CH CH CH 2.13 N CH CCl CH 2.14 CH CCl CCl CH 2.15 CH CF CH CH 2.16 CH CMe CH CH 2.17 CH N CH CH 2.18 CH CH CBr CH 2.19 CH CH CCF₃ CH 2.20 CH CH CEt CH 2.21 CH CH CF CH 2.22 CH CH CMe CH 2.23 CH CH C—OMe CH 2.24 CH CH CCl CCl 2.25 CH CH CH CF 2.26 CH CH CH C—OMe 2.27 CH CH CH N 2.28 Cl CH CH CH 2.29 CH CH CCN CH 2.30 CBr CH CCl CH

In some embodiments, A¹, A², A³, A⁴, A¹¹, A¹², A¹³ and A¹⁴ are selected from combinations 3.1-3.8 in the following table:

Combination A¹/A¹¹ A²/A¹² A³/A¹³ A⁴/A¹⁴ 1 CF CH CH CH 2 CH CBr CH CH 3 CH CCl CH CH 4 CF CCl CH CH 5 CH CH CH CBr 6 CH CH CCl CH 7 CH CF CF CH 8 CH CH CH CH

Of these, combinations 1, 3, 4, 6 and 7 may be preferred.

In further embodiments, A¹, A², A³, A⁴, A¹¹, A¹², A¹³ and A¹⁴ are selected from combinations 9-28 in the following table:

Combination A¹/A¹¹ A²/A¹² A³/A¹³ A⁴/A¹⁴  9 CF CH CF CH 10 CF CH CCl CH 11 CBr CH CH CH 12 CCl CH CH CH 13 N CH CCl CH 14 CH CCl CCl CH 15 CH CF CH CH 16 CH CMe CH CH 17 CH N CH CH 18 CH CH CBr CH 19 CH CH CCF₃ CH 20 CH CH CEt CH 21 CH CH CF CH 22 CH CH CMe CH 23 CH CH C—OMe CH 24 CH CH CCl CCl 25 CH CH CH CF 26 CH CH CH C—OMe 27 CH CH CH N 28 Cl CH CH CH 29 CH CH CCN CH 30 CBr CH CCl CH

Of these, combinations 9 and 10 may be preferred.

In some embodiments, combinations 1, 3, 6 and 9 may be preferred.

In some embodiments, combinations 1 and 10 may be preferred.

In some embodiments, combination 10 may be preferred.

In some embodiments, combination 1 may be preferred.

Y

In some embodiments, Y is (CH₂)_(n), where n is from 2 to 3. In some embodiments, Y is (CH₂)₂. In some embodiments, Y is (CH₂)₃. In some embodiments, Y is (CH₂)₄.

In other embodiments, Y is —CH₂—CH═CH—CH₂—.

R^(C1), R^(C3), R^(C4), R^(C11), R^(C13) and R^(C14)

In some embodiments, R^(C1), R^(C3), R^(C4), R^(C11), R^(C13) and R^(C14) are all H.

In other embodiments, two of R^(Cl), R^(C3) and R^(C4) and two of R^(C11), R^(C13) and R^(C14) are H, and the others are selected from the defined groups (except H).

In other embodiments, one of R^(C1), R^(C3) and R^(C4) and one of R^(C11), R^(C13) and R^(C14) is H, and the others are independently selected from the defined groups (except H).

In some embodiments, R^(C1), R^(C3) and R^(C4) are independently selected from H, Cl, F, Br, Me, OMe, cyano, CF₃ and CH₂OH. In some of these embodiments, R^(C1), R^(C3) and R^(C4) are independently selected from H, Cl, F, CF₃, OMe and CH₂OH. In further of these embodiments, R^(C1), R^(C3) and R^(C4) are independently selected from H, Cl and OMe. In other embodiments, R^(C1), R^(C3) and R^(C4) are independently selected from H, F and OMe. In further of these embodiments, R^(C1), R^(C3) and R^(C4) are independently selected from H and OMe.

In some embodiments, R^(C11), R^(C13) and R^(C14) are independently selected from H, Cl, F, Br, Me, OMe, cyano, CF₃ and CH₂OH. In some of these embodiments, R^(C11), R^(C13) and R^(C14) are independently selected from H, Cl, F, CF₃, OMe and CH₂OH. In further of these embodiments, R^(C11), R^(C13) and R^(C14) are independently selected from H, Cl and OMe. In other embodiments, R^(C1), R^(C3) and R^(C4) are independently selected from H, F and OMe. In further of these embodiments, R^(C11), R^(C13) and R^(C14) are independently selected from H and OMe.

In some embodiments, R^(C1) is OMe.

In some embodiments, R^(C11) is OMe.

In some embodiments, R^(C3) is H.

In some embodiments, R^(C13) is H.

In some embodiments, R^(C4) is H or F, preferably H.

In some embodiments, R^(C14) is H or F, preferably H.

In some embodiments, Fr and R^(C11) are OMe, R^(C3) and R^(C13) are H, and R^(C4) and R^(C14) are both either H or F, preferably H.

In some embodiments, R^(C1), R^(C3) and R^(C4) are selected from the following combinations 1-4:

Combination R^(C1) R^(C3) R^(C4) 1 H H H 2 H H F 3 OMe H H 4 OMe H F

In some embodiments, combination 3 and 4 may be preferred.

In some embodiments, combination 3 may be preferred.

In some embodiments, R^(C1), R^(C13) and R^(C14) are selected from the following combinations 1-4:

Combination R^(C11) R^(C13) R^(C14) 1 H H H 2 H H F 3 OMe H H 4 OMe H F

In some embodiments, combination 3 and 4 may be preferred.

In some embodiments, combination 3 may be preferred.

In some embodiments, R^(C1), R^(C3), R^(C4), R^(C11), R^(C13) and R^(C14) are selected from the following combinations 1-4:

Combination R^(C1)/R^(C11) R^(C3)/R^(C13) R^(C4)/R^(C14) 1 H H H 2 H H F 3 OMe H H 4 OMe H F

In some embodiments, combination 3 and 4 may be preferred.

In some embodiments, combination 3 may be preferred.

R¹ and R¹¹

In some embodiments, R¹ and R¹¹ are selected from C(═O)OH, C(═O)OCH₃, F, Br, tetrazolyl and oxo-oxadiazolyl.

In some embodiments, R¹ and R¹¹ are selected from C(═O)OH, tetrazolyl and oxo-oxadiazolyl.

In some embodiments, R¹ and R¹¹ are —C(═O)OH.

In other embodiments, R¹ and R¹¹ are tetrazolyl.

In other embodiments, R¹ and R¹¹ are oxo-oxadiazolyl, such as 5-oxo-4H-1,2,4-oxadiazol-3-yl and 2-oxo-3H-1,3,4-oxadiazol-5-yl.

In other embodiments, R¹ and R¹¹ are triazolyl.

In other embodiments, R¹ and R¹¹ are Br or F.

In some embodiments, R¹ and R¹¹ are not the same. In these embodiments, R^(C1) may be the same as R^(C11), R^(C3) may be the same as R^(C13) and R^(C14) may be the same as R^(C14), and A¹, A², A³ and A⁴ may be the same or different to A¹¹, A¹², A¹³ and A¹⁴, respectively. In some embodiments, R¹ is tetrazolyl and R¹¹ is C(═O)OH.

In some embodiments, R¹ and R¹¹ are C(═O)OR^(P1). In these embodiments, R^(P1) is preferably methyl.

Other Embodiments

In some embodiments, the compound is of formula (II):

wherein: Y is (CH₂)_(n), where n is from 2 to 4; R¹ and R¹¹ are selected from —C(═O)OH and tetrazolyl; A¹ is CR^(A) or N; A² is CR^(B) or N; A³ is CR^(C) or N; A⁴ is CR^(D) or N; where no more than two of A¹, A², A³, and A⁴ may be N; one, two or three of R^(A), R^(B), R^(C) and R^(D), (if present) are selected from H, F, Cl, Br, Me, CF₃, cyclopropyl, cyano, OMe, OEt, CH₂OH, CH₂OMe and OH; the remainder of R^(A), R^(B), R^(C) and R^(D), (if present) are H; A¹¹ is CR^(AA) or N; A¹² is CR^(BB) or N; A¹³ is CR^(CC) or N; A¹⁴ is CR^(DD) or N; where no more than two of A¹¹, A¹², A¹³ and A¹⁴ may be N; one, two or three of R^(AA), R^(BB), R^(CC) and R^(DD) (if present) are selected from H, F, Cl, Br, Me, CF₃, cyclopropyl, cyano, OMe, OEt, CH₂OH, CH₂OMe and OH; the remainder of R^(AA), R^(BB), R^(CC) and R^(DD), (if present) are H; R^(C1), R^(C3) and R^(C4) are independently selected from H, Cl, F, Br, Me, OMe, cyano, CF₃, CH₂OH, CH₂OMe, C₂₋₄ alkenyl and C₅heterocyclyl; R^(C11), R^(C13) and R^(C14) are independently selected from H, Cl, F, Br, Me, OMe, cyano, CF₃, CH₂OH, CH₂OMe, C₂₋₄alkenyl and C₅heterocyclyl.

EXAMPLES

The following examples are provided solely to illustrate the present invention and are not intended to limit the scope of the invention, as described herein.

Acronyms

For convenience, many chemical moieties are represented using well known abbreviations, including but not limited to, methyl (Me), ethyl (Et), n-propyl (nPr), isopropyl (iPr), n-butyl (nBu), tert-butyl (tBu), phenyl (Ph), benzyl (Bn), methoxy (MeO), ethoxy (EtO), trimethylsilyl (TMS), and acetyl (Ac).

For convenience, many chemical compounds are represented using well known abbreviations, including but not limited to, methanol (MeOH), deuterated methanol (MeOD-d₄ or CD₃OD) ethanol (EtOH), isopropanol (i-PrOH), ether or diethyl ether (Et₂O), ethyl acetate (EtOAc), acetic acid (AcOH), acetonitrile (MeCN or ACN), dichloromethane (methylene chloride, DCM), trifluoroacetic acid (TFA), dimethylformamide (DMF), tetrahydrofuran (THF), dimethylsulfoxide (DMSO), deuterated chloroform (CDCl₃), diethylamine (DEA), deuterated dimethylsulfoxide (DMSO-d₆), N-ethyl-N′-(3-dimethylaminopropyl)carbodiimide hydrochloride (EDCl.HCl), meta-chloroperoxybenzoic acid (mCPBA), 1,1′-bis(diphenylphosphino)ferrocene (dppf), tert-butyloxycarbonyl (Boc, BOC), 2-(trimethylsilyl)ethoxymethyl (SEM), triethylamine (Et₃N or TEA), 2-(1H-7-azabenzotriazol-1-yl)-1,1,3,3-tetramethyluronium hexafluorophosphate (HATU), 4-dimethylaminopyridine (DMAP), N,N-diisopropylethylamine (DIPEA or DIEA), 1,1′-bis(diphenylphosphino)ferrocene dichloropalladium (II) (PdCl₂(dppf)), trans-dichlorobis(triphenylphosphine)palladium(II) (PdCl₂(PPh₃)₂), tris(dibenzylideneacetone) dipalladium(0) (Pd₂(dba)₃), tetrakis(triphenylphosphine)palladium(0) (Pd(PPh₃)₄), propylphosphonic anhydride (T3P), hexamethylphosphoramide (HMPA), 1,2-dichloroethane (DCE), chromium(VI) oxide (CrO₃), n-bromosuccinimide (NBS), potassium hydroxide (KOH), benzoyl peroxide (BPO), carbon tetrachloride (CCl₄), petroleum ether (Pet. Ether), potassium carbonate (K₂CO₃), sodium sulfate (Na₂SO₄), lithium diisopropylamine (LDA), azobisisobutyronitrile (AIBN), N-methylmorpholine N-oxide (NMO), benzoyl peroxide (BPO), 1,1′-carbonyldiimidazole (CDI) and 1-hydroxybenzotriazole (HOBt).

In addition, TLC refers to thin layer chromatography.

General Experimental Details

Unless otherwise stated the following generalisations apply. ¹H NMR spectra were recorded on a Bruker AVANCE III (400 MHz). The multiplicity of a signal is designated by the following abbreviations: s, singlet; d, doublet; t, triplet; q, quartet; dd, doublet of doublets; dt, doublet of triplets; tt, triplet of triplets; td, triplet of doublets; ddd, doublet of doublet of doublets; br, broad; m, multiplet. All observed coupling constants, J, are reported in Hertz. Exchangeable protons are not always observed.

LCMS data was generated using the conditions described below. Chlorine isotopes are reported as ³⁵Cl, Bromine isotopes are reported as either ⁷⁹Br or ⁸¹Br or both ⁷⁹Br/⁶¹Br.

LC-MS Method A (LCMS-A):

Equipment Information

LC model: Agilent 1200

(Pump type: Binary Pump, Detector type: DAD)

MS model: Agilent G6110A Quadrupole

Parameters of LCMS

LC: Column: Xbridge-C18, 2.5 μm, 2.1×30 mm

-   -   Column temperature: 30° C.     -   Acquisition of wavelength: 214 nm, 254 nm     -   Mobile phase: A: 0.07% HCOOH aqueous solution, B: MeOH

MS: Ion source: ES+ (or ES−) MS range: 50˜900 m/z

-   -   Fragmentor: 60 Drying gas flow: 10 L/min     -   Nebulizer pressure: 35 psi Drying gas temperature: 350° C.     -   Vcap: 3.5 kV

Gradient Table: Flow (mL/min) T (min) A (%) B (%) 0.5 0.0 70 30 0.5 0.2 70 30 0.5 1.8 5 95 0.5 2.4 5 95 0.5 2.6 70 30 0.5 3.5 70 30

Sample Preparation

The sample was dissolved in methanol, the concentration about 0.11˜1 mg/mL, then filtered through a 0.22 μm syringe filter. (Injection volume: 1˜10 μL)

LC-MS Method B (LCMS-B):

Equipment Information

LC model: Agilent 1200

(Pump type: Binary Pump, Detector type: DAD)

MS model: Agilent G6110A Quadrupole

Parameters of LCMS

LC: Column: Xbridge-C18, 2.5 μm, 2.1×30 mm

-   -   Column temperature: 30° C.     -   Acquisition of wavelength: 214 nm, 254 nm     -   Mobile phase: A: 0.07% HCOOH aqueous solution, B: MeOH

MS: Ion source: ES+ (or ES−) MS range: 50˜900 m/z

-   -   Fragmentor: 60 Drying gas flow: 10 L/min     -   Nebulizer pressure: 35 psi Drying gas temperature: 350° C.     -   Vcap: 3.5 kV

Gradient Table: Flow (mL/min) T (min) A(%) B (%) 0.5 0.0 70 30 0.5 0.3 70 30 0.5 0.6 50 50 0.5 0.9 40 60 0.5 1.2 30 70 0.5 3.2 5 95 0.5 3.5 5 95 0.5 4.0 70 30 0.5 5.0 70 30

Sample Preparation

The sample was dissolved in methanol, the concentration about 0.11˜1 mg/mL, then filtered through a 0.22 μm syringe filter. (Injection volume: 1˜10 μL)

LC-MS Method C (LCMS-C):

Equipment Information

LC model: Waters 2695 alliance

(Pump type: Quaternary Pump, Detector: 2996 Photodiode Array Detector)

MS model: Micromass ZQ

Parameters of LCMS

LC: Column: Xbridge-C18, 3.5 μm, 2.1×50 mm

-   -   Column temperature: 30° C.     -   Acquisition of wavelength: 214 nm, 254 nm     -   Mobile phase: A: 0.07% HCOOH aqueous solution, B: MeOH

MS: Ion source: ES+ (or ES−) MS range: 50˜900 m/z

-   -   Capillary: 3 kV Cone: 3 V Extractor: 3 V     -   Drying gas flow: 600 L/hr Cone: 50 L/hr     -   Desolvation temperature: 300° C.     -   Source temperature: 100° C.

Gradient Table: Flow (mL/min) T (min) A (%) B (%) 0.3 0.0 80 20 0.3 0.5 80 20 0.3 0.8 50 50 0.3 1.2 35 65 0.3 2.0 20 80 0.3 4.0  5 95 0.3 5.0  5 95 0.3 5.8 15 85 0.3 6.2 80 20 0.3 8.0 80 20

Sample Preparation

The sample was dissolved in methanol, the concentration about 0.11˜1 mg/mL, then filtered through a 0.22 μm syringe filter. (Injection volume: 1˜10 μL)

Other Chromatography

Analytical thin-layer chromatography was performed on Merck silica gel 60 F254 aluminium-backed plates which were visualised using fluorescence quenching under UV light or a basic KMnO₄ dip or Ninhydrin dip.

Preparative thin-layer chromatography (prep TLC) was performed using Tklst (China), grand grade: (HPTLC): 8±2 μm>80%; (TLC): 10-40 μm. Type: GF254. Compounds were visualised by UV (254 nm).

Column chromatography was performed using Tklst (China), grand grade, 100-200 meshes silica gel.

Where necessary, anhydrous solvents were purchased from Sigma-Aldrich or dried using conventional methods. Solutions of inorganic acids or bases were made up as aqueous solutions unless stated otherwise.

Solutions of hydrogen chloride, sodium hydroxide, potassium carbonate and sodium bicarbonate are aqueous, unless otherwise stated.

Intermediate Preparations (i) 4,4′-(Ethane-1,2-diylbis(azanediyl))bis(3-aminobenzamide) (I4)

(a) tert-Butyl (2-((4-carbamoyl-2-nitrophenyl)amino)ethyl)carbamate (I1)

A suspension of 4-chloro-3-nitrobenzamide (8.0 g, 39.9 mmol), tert-butyl (2-aminoethyl)carbamate (6.40 g, 39.9 mmol) and Et₃N (8.1 g, 79.8 mmol) in NMP (80 mL) was heated at 150° C. under N₂ overnight. Water (500 mL) was added followed by Pet. Ether/EtOAc (5:1, 300 mL) and the resulting precipitate was collected by filtration to give the title compound (12.0 g, 92%) as a yellow solid. LCMS-B: rt 3.2 min, m/z 347.1 [M+Na]⁺. ¹H NMR (400 MHz, DMSO-d₆) δ 8.65 (s, 1H), 8.44 (t, J=5.8 Hz, 1H), 8.05-7.92 (m, 2H), 7.27 (s, 1H), 7.14 (d, J=9.1 Hz, 1H), 7.04 (t, J=5.8 Hz, 1H), 3.46-3.45 (m, 2H), 3.20-3.19 (m, 2H), 1.35 (s, 9H).

(b) 4-((2-Aminoethyl)amino)-3-nitrobenzamide hydrochloride (I2)

A mixture of tert-butyl (2-((4-carbamoyl-2-nitrophenyl)amino)ethyl)carbamate (I1) (12.0 g, 37.0 mmol) and a 5.5 M HCl in dioxane solution (200 mL) was stirred at room temperature overnight. The solvent was then removed under reduced pressure to give the title compound (8.0 g, 96%) as a yellow solid. LCMS-B: rt 1.2 min, m/z 225.1 [M+H]⁺. ¹H NMR (400 MHz, DMSO-d₆) δ 8.65 (d, J=2.1 Hz, 1H), 8.42 (s, 1H), 8.25 (s, 2H), 8.05 (dd, J=9.0, 2.2 Hz, 1H), 7.25 (d, J=9.0 Hz, 1H), 5.62 (br s, 3H), 3.76-3.74 (m, 2H), 3.01-2.97 (m, 2H).

(c) 4,4′-(Ethane-1,2-diylbis(azanediyl))bis(3-nitrobenzamide) (I3)

A suspension of 4-((2-aminoethyl)amino)-3-nitrobenzamide hydrochloride (I2) (7.6 g, 33.9 mmol), 4-chloro-3-nitrobenzamide (6.80 g, 33.9 mmol) and Et₃N (20.6 g, 203 mmol) in NMP (80 mL) was heated at 150° C. under N₂ overnight. Additional 4-((2-aminoethyl)amino)-3-nitrobenzamide hydrochloride (12) (800 mg, 3.57 mmol) and Et₃N (3.0 g, 29.6 mmol) were added and the mixture was heated at 150° C. under N₂ overnight. The mixture was allowed to cool to room temperature, diluted with Pet. Ether/EtOAc (1:1, v/v, 500 mL) and the resulting precipitate was collected by filtration to give the title compound (12.0 g, 91%) as a yellow solid. LCMS-B: rt 1.3 min, m/z 389.0 [M+H]⁺. ¹H NMR (400 MHz, DMSO-d₆) δ 8.62 (s, 4H), 7.99-7.95 (m, 4H), 7.30-7.28 (m, 4H), 3.70-3.64 (m, 4H).

(d) 4,4′-(Ethane-1,2-diylbis(azanediyl))bis(3-aminobenzamide) (I4)

A suspension of 4,4′-(ethane-1,2-diylbis(azanediyl))bis(3-nitrobenzamide) (I3) (12 g, 30.9 mmol), and 10% Pd/C (2.0 g) in DMF (200 mL) was heated at 100° C. under a H2 atmosphere overnight. The mixture was filtered and the filtrate was concentrated under reduced pressure to give the title compound (9.0 g, 90%) as a black solid. LCMS-B: rt 0.3 min, m/z 329.1 [M+H]⁺. ¹H NMR (400 MHz, DMSO-d₆) δ 7.44 (s, 2H), 7.13 (d, J=8.2 Hz, 2H), 7.11 (s, 2H), 6.74 (s, 2H), 6.46 (d, J=8.0 Hz, 2H), 5.05 (s, 2H), 4.58 (s, 4H), 3.41-3.36 (m, 4H).

(ii) 4,4′-(Ethane-1,2-diylbis(azanediyl))bis(3-amino-2-methoxybenzamide) I13

(a) Methyl 4-fluoro-2-hydroxybenzoate (I5)

A mixture of 4-fluoro-2-hydroxybenzoic acid (50.0 g, 320.3 mmol) and concentrated H₂SO₄ (40 mL, 672.7 mmol) in MeOH (600 mL) was heated at reflux under N₂ for 16 h. The mixture was poured into water (500 mL) and extracted with EtOAc (500 mL×3). The combined organic extracts were washed with water (500 mL×2), brine (500 mL), dried over Na₂SO₄, filtered and concentrated under reduced pressure. The residue was purified by silica gel chromatography (Pet. Ether/EtOAc=100:1 to 20:1) to give the title compound (50.0 g, 91%) as a white solid. LCMS-A (ES-API): rt 1.80 min, m/z 171.1 [M+H]⁺. ¹H NMR (400 MHz, DMSO-d₆) δ 10.8 (s, 1H), 7.83 (dd, J=8.9, 6.8 Hz, 1H), 6.90-6.70 (m, 2H), 3.88 (s, 3H).

(b) Methyl 4-fluoro-2-hydroxy-3-nitrobenzoate (I6)

To a solution of methyl 4-fluoro-2-hydroxybenzoate I5 (50.0 g, 294.0 mmol) in concentrated H₂SO₄ (100 mL) at 0° C. under N₂ was added concentrated HNO₃ (20 mL, 382.1 mmol) dropwise and the mixture was allowed to warm to room temperature and stirred for 16 h. The mixture was slowly poured into water (2.0 L) and extracted with EtOAc (1.5 L×3). The combined organic extracts were washed water (1.5 L×2), brine (1.5 L), dried over Na₂SO₄, filtered and concentrated under reduced pressure. The residue was purified by silica gel chromatography (100% Pet. Ether) to give a mixture of the title compound and an uncharacterised regioisomer (50 g, ˜3.75:1 uncharacterised regioisomer/title compound). The mixture was used in the next step without further purification or characterisation. LCMS-A (ES-API): rt 1.57 min (minor) m/z 216.0 [M+H]+ and rt 1.68 min (major) m/z 216.1 [M+H]⁺.

(c) Methyl 4-fluoro-2-methoxy-3-nitrobenzoate (I7)

A mixture of methyl 4-fluoro-2-hydroxy-3-nitrobenzoate (containing a major, uncharacterised regioisomer) I6 (50.0 g), CH₃I (99.0 g, 697 mmol) and K₂CO₃ (64.1 g, 465 mml) in DMF (200 mL) was stirred at room temperature under N₂ for 16 h. The mixture was poured into water (1.0 L), extracted with EtOAc (1.0 L×3) and the combined organic extracts were washed water (1.0 L×2), brine (1.0 L), dried over Na₂SO₄, filtered and concentrated under reduced pressure. The residue was purified by silica gel chromatography (Pet. Ether/EtOAc=100:1 to 40:1) and the fractions containing the minor component were collected to give the title compound (14.0 g, 21% over two steps) as a white solid. LCMS-C (ES-API): rt 3.53 min, m/z 230.0 [M+H]⁺. ¹H NMR (400 MHz, DMSO-d₆) δ 8.12 (dd, J=9.1, 6.5 Hz, 1H), 7.50 (t, J=9.0 Hz, 1H), 3.93 (s, 3H), 3.91 (s, 3H).

(d) Methyl 4-((2-((tert-butoxycarbonyl)amino)ethyl)amino)-2-methoxy-3-nitrobenzoate (I8)

A mixture of methyl 4-fluoro-2-methoxy-3-nitrobenzoate I7 (6.0 g, 26.2 mmol), tert-butyl (2-aminoethyl) carbamate (4.2 g, 26.2 mmol) and Et₃N (5.3 g, 52.4 mmol) in NMP (100 mL) was heated at 80° C. under N₂ for 16 h. The mixture was poured into water (500 mL), extracted with EtOAc (500 mL×3) and the combined organic extracts were washed with water (500 mL×2), brine (500 mL), dried over Na₂SO₄, filtered and concentrated under reduced pressure. The residue was purified by silica gel chromatography (Pet. Ether/EtOAc=100:1 to 10:1) to give the title compound (9.5 g, 98%) as a yellow solid. LCMS-C (ES-API): rt 4.43 min, m/z 392.1 [M+Na]⁺. ¹H NMR (400 MHz, DMSO-d₆) δ 7.79 (d, J=9.2 Hz, 1H), 6.96 (t, J=5.5 Hz, 1H), 6.74 (d, J=9.2 Hz, 1H), 6.70 (t, J=5.7 Hz, 1H), 3.79 (s, 6H), 3.27-3.21 (m, 2H), 3.11-3.06 (m, 2H), 1.37 (s, 9H).

(e) Methyl 4-((2-aminoethyl)amino)-2-methoxy-3-nitrobenzoate hydrochloride (I9)

A mixture of methyl 4-((2-((tert-butoxycarbonyl)amino)ethyl)amino)-2-methoxy-3-nitrobenzoate I8 (9.5 g, 25.7 mmol) and a 3 M HCl in dioxane solution (200 mL) was stirred at room temperature under N₂ for 16 h. The mixture was concentrated under reduced pressure to give the title compound (6.0 g, 87%) as a white solid. LCMS-C (ES-API): rt 0.82 min, m/z 270.0 [M+H]⁺. ¹H NMR (400 MHz, DMSO-d₆) δ 7.81 (dd, J=9.2, 3.8 Hz, 1H), 6.82-6.71 (m, 2H), 3.79 (s, 3H), 3.75 (s, 3H), 3.31-3.28 (m, 4H).

(f) Dimethyl 4,4′-(ethane-1,2-diylbis(azanediyl))bis(2-methoxy-3-nitrobenzoate) (I10)

A mixture of methyl 4-((2-aminoethyl)amino)-2-methoxy-3-nitrobenzoate hydrochloride I9 (5.5 g, 18.0 mmol), methyl 4-fluoro-2-methoxy-3-nitrobenzoate 17 (4.68 g, 20.4 mmol) and Et₃N (6.2 g, 61.3 mmol) in NMP (100 mL) was heated at 80° C. under N₂ for 16 h. The mixture was poured into water (600 mL) and the resulting precipitate was collected by filtration to give the title compound (8.0 g, 82%) as a yellow solid. LCMS-C (ES-API): rt 4.75 min, m/z 477.1 [M−H]⁻. ¹H NMR (400 MHz, DMSO-d₆) δ 7.76 (d, J=9.2 Hz, 2H), 6.77-6.74 (m, 4H), 3.78 (s, 6H), 3.77 (s, 6H), 3.40 (br s, 4H).

(g) 4,4′-(Ethane-1,2-diylbis(azanediyl))bis(2-methoxy-3-nitrobenzoic acid) (I11)

A mixture of dimethyl 4,4′-(ethane-1,2-diylbis(azanediyl))bis(2-methoxy-3-nitrobenzoate) I10 (4.0 g, 8.4 mmol) and NaOH (2.0 g, 50.2 mmol) in EtOH/water (50 mL/50 mL) was heated at 80° C. for 16 h. Most of the EtOH was removed under reduced pressure and the residue was diluted with water (50 mL) and acidified to pH 4-5 with a 2 M aqueous HCl solution. The resulting precipitate was collected by filtration to give the title compound (3.5 g, 93%) as a yellow solid. LCMS-C (ES-API): rt 3.45 min, m/z 473.1 [M+Na]⁺. ¹H NMR (400 MHz, DMSO-d₆) δ 12.6 (br s, 2H), 7.77 (d, J=9.1 Hz, 2H), 6.74 (d, J=9.3 Hz, 2H), 6.69 (t, J=5.5 Hz, 2H), 3.78 (s, 6H), 3.44 (br s, 4H).

(h) 4,4′-(Ethane-1,2-diylbis(azanediyl))bis(2-methoxy-3-nitrobenzamide) (I12)

A mixture of 4,4′-(ethane-1,2-diylbis(azanediyl))bis(2-methoxy-3-nitrobenzoic acid) I11 (3.4 g, 7.6 mmol), NH₄Cl (2.4 g, 45.3 mmol), HOBt (3.1 g, 22.7 mmol), EDCl.HCl (4.3 g, 22.7 mmol) and DIPEA (5.9 g, 45.3 mmol) in DMF (300 mL) was stirred at room temperature under N₂ for 16 h. The mixture was poured into water (600 mL) and the resulting precipitate was collected by filtration to give the title compound (2.4 g, 71%) as a yellow solid. LCMS-C (ES-API): rt 3.02 min, m/z 471.1 [M+Na]⁺. ¹H NMR (400 MHz, DMSO-d₆) δ 7.63 (d, J=9.0 Hz, 2H), 7.42 (br s, 4H), 6.77 (d, J=9.1 Hz, 2H), 6.60 (t, J=4.8 Hz, 2H), 3.77 (s, 6H), 3.39-3.36 (m, 4H).

(i) 4,4′-(Ethane-1,2-diylbis(azanediyl))bis(3-amino-2-methoxybenzamide) (I13)

A mixture of 4,4′-(ethane-1,2-diylbis(azanediyl))bis(2-methoxy-3-nitrobenzamide) I12 (1.0 g, 2.3 mmol) and 10% Pd/C (200 mg) in MeOH (150 mL) was stirred at room temperature under a H₂ atmosphere (3 MPa) for 3 days. The mixture was filtered and the filtrate was concentrated under reduced pressure give the title compound (120 mg). The filter cake was washed with DMSO and the filtrate was freeze-dried to give additional title compound (680 mg). The isolated solids were combined to give the title compound (800 mg, 92%) as a brown solid. LCMS-B (ES-API): rt 1.09 min, m/z 388.8 [M+H]⁺. ¹H NMR (400 MHz, DMSO-d₆) δ 7.39 (br s, 2H), 7.11 (d, J=8.4 Hz, 2H), 7.10 (br s, 2H), 6.37 (d, J=8.5 Hz, 2H), 5.23 (br s, 2H), 4.42 (br s, 4H), 3.66 (s, 6H), 3.35 (br s, 4H).

Examples Example 1: 6,6′-(Ethane-1,2-diylbis(5-carbamoyl-4-methoxy-1H-benzo[d]imidazole-1,2-diyl))bis(3-chlorobenzoic acid)

(a) Methyl 2-(bromomethyl)-5-chlorobenzoate (A1)

To a solution of methyl 5-chloro-2-methylbenzoate (5.0 g, 27.1 mmol) in 0014 (50 mL) was added NBS (5.3 g, 29.8 mmol) and AIBN (2.2 g, 13.5 mmol) and the mixture was heated at reflux under N₂ for 4 h. The mixture was poured into water, extracted with EtOAc and the combined organic extracts were washed with brine, dried over Na₂SO₄, filtered and concentrated under reduced pressure. The residue was purified by column chromatography (Pet. Ether) to give the title compound (3.0 g, 34%) as a colorless oil. ¹H NMR (400 MHz, DMSO-d₆) δ 7.85 (d, J=2.2 Hz, 1H), 7.69-7.62 (m, 2H), 4.99 (s, 2H), 3.88 (s, 3H).

(b) Methyl 5-chloro-2-formylbenzoate (A2)

To a solution of methyl 2-(bromomethyl)-5-chlorobenzoate A1 (480 mg, 1.83 mmol) in THF (7.0 mL) was added NMO (429 mg, 3.66 mmol) and the mixture was heated at 100° C. under N₂ for 16 h. The mixture was poured into water (30 mL), extracted with EtOAc (30 mL×3) and the combined organic extracts were washed brine, dried over Na₂SO₄, filtered and concentrated under reduced pressure. The residue was purified by column chromatography (Pet. Ether/EtOAc=1:0 to 250:1) to give the title compound (170 mg, 47%) as a white solid. LCMS-B (ES-API): rt 3.68 min, m/z 198.8 [M+H]⁺. ¹H NMR (400 MHz, DMSO-d₆) δ 10.3 (s, 1H), 7.93-7.83 (m, 3H), 3.92 (s, 3H).

(c) Dimethyl 6,6′-(ethane-1,2-diylbis(5-carbamoyl-4-methoxy-1H-benzo[d]imidazole-1,2-diyl))bis(3-chlorobenzoate) (A3)

A mixture of methyl 5-chloro-2-formylbenzoate A2 (83 mg, 0.42 mmol), 4,4′-(ethane-1,2-diylbis(azanediyl))bis(3-amino-2-methoxybenzamide) 113 (80 mg, 0.21 mmol) and NaHSO₃ (44 mg, 0.42 mmol) in DMF (4 mL) was heated at 100° C. under N₂ for 16 h. The mixture was diluted with water until a precipitate formed, which was collected by filtration to give the title compound (130 mg, 83%) as a brown solid. LCMS-B (ES-API): rt 3.95 min, m/z 745.4 [M+H]⁺. ¹H NMR (400 MHz, DMSO-d₆) δ 7.80 (d, J=2.3 Hz, 2H), 7.71 (s, 2H), 7.68 (d, J=8.6 Hz, 2H), 7.45 (s, 2H), 7.28 (dd, J=8.2, 2.3 Hz, 2H), 7.06 (d, J=8.6 Hz, 2H), 6.79 (d, J=8.2 Hz, 2H), 4.36 (s, 6H), 4.25 (s, 4H), 3.61 (s, 6H).

(d) 6,6′-(Ethane-1,2-diylbis(5-carbamoyl-4-methoxy-1H-benzo[d]imidazole-1,2-diyl))bis(3-chlorobenzoic acid) (1)

A mixture of dimethyl 6,6′-(ethane-1,2-diylbis(5-carbamoyl-4-methoxy-1H-benzo[d]imidazole-1,2-diyl))bis(3-chlorobenzoate) A3 (120 mg, 0.16 mmol) and NaOH (39 mg, 0.96 mmol) in EtOH/H₂O (5 mL/5 mL) was heated at 80° C. for 3 h. Most of the EtOH was removed under reduced pressure and the residue was adjusted to pH 4-5 with a 1 M aqueous HCl solution. The resulting precipitate was collected by filtration and purified by trituration with DCM/MeOH (10:1) to give the title compound (44 mg, 38%) as a brown solid. LCMS-B (ES-API): rt 2.66 min, m/z 717.3 [M+H]⁺. ¹H NMR (400 MHz, DMSO-d₆) δ 7.80 (d, J=2.3 Hz, 2H), 7.71 (s, 2H), 7.64 (d, J=8.6 Hz, 2H), 7.44 (s, 2H), 7.14 (dd, J=8.4, 1.2 Hz, 2H), 6.94 (d, J=8.6 Hz, 2H), 6.63 (d, J=8.2 Hz, 2H), 4.39 (s, 6H), 4.27 (s, 4H).

Example 2—2,2′-(Ethane-1,2-diylbis(5-carbamoyl-1H-benzo[d]imidazole-1,2-diyl))dibenzoic acid (2)

(a) Dimethyl 2,2′-(ethane-1,2-diylbis(5-carbamoyl-1H-benzo[d]imidazole-1,2-diyl))dibenzoate (A4)

A solution of 4,4′-(ethane-1,2-diylbis(azanediyl))bis(3-aminobenzamide) 14 (500 mg 1.52 mmol), methyl 2-formylbenzoate (499 mg 3.04 mmol) and NaHSO₃ (316 mg 3.04 mmol) in DMF (30 mL) was heated at 120° C. under N₂ overnight. The resulting precipitate was collected by filtration, washed with water and DCM and dried under reduced pressure to give the title compound (100 mg, 26%) as a grey solid. LCMS-B (ES-API): rt 2.59 min; m/z 617.2 [M+H]⁺. ¹H NMR (400 MHz, DMSO-d₆) δ 8.18 (d, J=1.0 Hz, 2H), 8.01 (br s, 2H), 7.89 (dd, J=7.9, 1.3 Hz, 2H), 7.65 (dd, J=8.5, 1.6 Hz, 2H), 7.54-7.43 (m, 2H), 7.28 (br s, 2H), 7.22 (d, J=8.5 Hz, 2H), 7.21-7.16 (m, 2H), 6.52 (d, J=7.5 Hz, 2H), 4.26 (s, 4H), 3.53 (m, 6H).

(b) 2,2′-(Ethane-1,2-diylbis(5-carbamoyl-1H-benzo[d]imidazole-1,2-diyl))dibenzoic acid (2)

A mixture of 2,2′-(ethane-1,2-diylbis(5-carbamoyl-1H-benzo[d]imidazole-1,2-diyl))dibenzoate A4 (120 mg 0.19 mmol) and NaOH (47 mg 1.17 mmol) in EtOH/water (10 mL/2 mL) was heated at 50° C. under N₂ overnight. The solvent was removed under reduced pressure and the residue was diluted with water (3 mL) and acidified to pH 4-5 with 1 M aqueous HCl solution. The resulting precipitate was collected by filtration, washed with water and dried at 60° C. to give the title compound (25 mg, 22%) as a black solid. LCMS-B (ES-API): rt 0.33 min; m/z 589.2 [M+H]⁺. ¹H NMR (400 MHz, DMSO-d₆) δ 8.22 (s, 2H), 8.08 (br s, 2H), 7.91 (d, J=7.5 Hz, 2H), 7.67 (d, J=8.5 Hz, 2H), 7.52 (t, J=7.4 Hz, 2H), 7.41 (br s, 2H), 7.36-7.28 (m, 2H), 7.21-7.28 (m, 2H), 6.74-6.59 (m, 2H), 4.42 (s, 4H).

Example 3—2,2′-(Ethane-1,2-diylbis(5-carbamoyl-1H-benzo[d]imidazole-1,2-diyl)bis(4-bromobenzoic acid) (3)

(a) Methyl 4-bromo-2-formylbenzoate (A5)

To a solution of methyl 4-bromo-2-(bromomethyl)benzoate (2.0 g, 6.5 mmol) in acetonitrile (31 mL) at 0° C. was added NMO (50% aqueous solution, 4.6 g, 19.7 mmol) and the mixture was stirred at 0° C. overnight. The mixture was diluted with water, extracted with EtOAc and the organic layer was washed with a saturated aqueous NaHCO₃ solution, dried over Na₂SO₄, filtered and concentrated under reduced pressure. The residue was purified by silica gel chromatography (Pet. ether/EtOAc=50:1) to give the title compound (622 mg, 39%) as a white solid. LCMS-B (ES-API): rt 3.93 min, m/z 242.9 [M+H]⁺. ¹H NMR (400 MHz, DMSO-d₆) δ 10.4 (s, 1H), 8.01-7.95 (m, 2H), 7.85 (d, J=8.2 Hz, 1H), 3.90 (s, 3H).

(b) Dimethyl 2,2′-(ethane-1,2-diylbis(5-carbamoyl-1H-benzo[d]imidazole-1,2-diyl))bis(4-bromobenzoate) (A6)

To a solution of methyl 4-bromo-2-formylbenzoate A5 (200 mg, 0.82 mmol) and 4,4′-(ethane-1,2-diylbis(azanediyl))bis(3-aminobenzamide) 14 (135 mg, 0.41 mmol) in DMF (9 mL) was added NaHSO₃ (85 mg, 0.82 mmol) and the mixture was heated at 120° C. under N₂ overnight. Water was added and the mixture was extracted with EtOAc. The combined organic extracts were washed with water, brine, dried over Na₂SO₄, filtered and concentrated under reduced pressure to give the title compound (165 mg, 52%) as a yellow solid. LCMS-B (ES-API): rt 2.99 min, m/z 773.0/775.0 [M+H]⁺. ¹H NMR (400 MHz, DMSO-d₆) δ 8.25 (s, 2H), 7.97 (br s, 2H), 7.82 (d, J=8.4 Hz, 2H), 7.76-7.73 (m, 4H), 7.32 (d, J=8.5 Hz, 2H), 7.27 (br s, 2H), 6.82 (s, 2H), 4.31 (s, 4H), 3.58 (s, 6H).

(c) 2,2′-(Ethane-1,2-diylbis(5-carbamoyl-1H-benzo[d]imidazole-1,2-diyl))bis(4-bromobenzoic acid) (3)

To a solution of dimethyl 2,2′-(ethane-1,2-diylbis(5-carbamoyl-1H-benzo[d]imidazole-1,2-diyl))bis(4-bromobenzoate) A6 (100 mg, 0.13 mmol) in EtOH (6 mL) was added a solution of NaOH (31 mg, 0.78 mmol) in water (4 mL) and the mixture was stirred at room temperature overnight. Most of the EtOH was removed under reduced pressure and the residue was diluted with water, acidified to pH 3-4 with a 2 M aqueous HCl solution and the resulting precipitate was collected by filtration and dried under reduced pressure to give the title compound (17 mg, 18%) as a yellow solid. LCMS-B (ES-API): rt 2.48 min, m/z 745.0/747.0 [M+H]⁺. 1H NMR (400 MHz, DMSO-d₆) δ 8.26 (s, 2H), 8.00 (br s, 2H), 7.83 (d, J=8.0 Hz, 2H), 7.75-7.65 (m, 4H), 7.37-7.21 (m, 4H), 6.79 (br s, 2H), 4.39 (s, 4H).

Example 4—2,2′-(Ethane-1,2-diylbis(5-carbamoyl-4-methoxy-1H-benzo[d]imidazole-1,2-diyl))dibenzoic acid (4)

(a) Dimethyl 2,2′-(ethane-1,2-diylbis(5-carbamoyl-4-methoxy-1H-benzo[d]imidazole-1,2-diyl))dibenzoate (A7)

A mixture of methyl 2-formylbenzoate (197 mg, 1.20 mmol), 4,4′-(ethane-1,2-diylbis(azanediyl))bis(3-amino-2-methoxybenzamide) 113 and NaHSO₃ (52 mg, 0.50 mmol) in DMF (20 mL) was heated at 120° C. under a N₂ atmosphere overnight. The mixture was diluted with water and partitioned against EtOAc, resulting in formation of a precipitate. The precipitate was collected by filtration and triturated with DCM/MeOH=20/1 then dried under reduced pressure to give title compound (115 mg, 14%) as a white solid. LCMS-C (ES-API): rt 3.1 min, m/z 677.4 [M+H]⁺. ¹H NMR (400 MHz, DMSO-d₆) δ 7.91 (d, J=7.9 Hz, 2H), 7.70 (br s, 2H), 7.61 (d, J=8.6 Hz, 2H), 7.56-7.51 (m, 2H), 7.46 (br s, 2H), 7.34 (t, J=8.4 Hz, 2H), 6.86 (d, J=8.6 Hz, 2H), 6.70 (d, J=7.2 Hz, 2H), 4.36 (s, 6H), 4.22 (s, 4H), 3.56 (s, 6H).

(b) 2,2′-(Ethane-1,2-diylbis(5-carbamoyl-4-methoxy-1H-benzo[d]imidazole-1,2-diyl))dibenzoic acid (4)

A mixture of dimethyl 2,2′-(ethane-1,2-diylbis(5-carbamoyl-4-methoxy-1H-benzo[d]imidazole-1,2-diyl))dibenzoate A7 (70 mg, 0.10 mmol) and NaOH (40.0 mg, 1.00 mmol) in MeOH/water (10 mL/1 mL) was stirred at room temperature overnight. Most of the MeOH was removed under reduced pressure and the residue was acidified to pH 5 with a 1 M aqueous HCl solution. The resulting precipitate was collected by filtration and dried under reduced pressure to give the title compound (16 mg, 25%) as a grey solid. LCMS-C (ES-API): rt 2.3 min, m/z 649.1 [M+H]⁺. 1H NMR (400 MHz, DMSO-d₆) δ 7.94 (d, J=7.9 Hz, 2H), 7.70 (br s, 2H), 7.56 (d, J=8.6 Hz, 2H), 7.51 (t, J=7.8 Hz, 2H), 7.45 (br s, 2H), 7.27 (t, J=7.5 Hz, 2H), 6.69 (d, J=8.7 Hz, 2H), 6.54 (d, J=7.6 Hz, 2H), 4.37 (s, 6H), 4.18 (s, 4H).

Example 5—2-(2-(1H-Tetrazol-5-yl)phenyl)-1-(2-(2-(2-(2H-tetrazol-5-yl)phenyl)-5-carbamoyl-1H-benzo[d]imidazol-1-yl)ethyl)-1H-benzo[d]imidazole-5-carboxamide (5)

(a) 2-(1H-Tetrazol-5-yl)benzaldehyde (A8)

To a solution of 5-phenyl-1H-tetrazole (5.0 g, 34.2 mmol) in THF (100 mL) at −78° C. under nitrogen was added s-butyllithium (1.3 M, 54.0 mL, 69.0 mmol) dropwise and the mixture was stirred at −78° C. for 30 min. DMF (20 mL, 300 mmol) was added dropwise and the mixture was allowed to warm to room temperature and stirred for 10 min. The reaction was quenched with a 1 M aqueous HCl solution (100 mL) and the mixture was extracted with EtOAc (300 mL×3). The combined organic extracts were washed with brine, dried over MgSO₄, filtered and concentrated under reduced pressure. The residue was purified by silica gel chromatography (Pet. ether/EtOAc=5:1-4:1) to give the title compound (4.0 g, 67%) as a white solid. LCMS-B (ES-API): rt 1.03 min; m/z 175.0 [M+H]⁺.

(b) 2-(2-(1H-Tetrazol-5-yl)phenyl)-1-(2-(2-(2-(2H-tetrazol-5-yl)phenyl)-5-carbamoyl-1H-benzo[d]imidazol-1-yl)ethyl)-1H-benzo[d]imidazole-5-carboxamide (5)

A mixture of 2-(1H-tetrazol-5-yl)benzaldehyde A8 (500 mg, 2.88 mmol) 4,4′-(ethane-1,2-diylbis(azanediyl))bis(3-aminobenzamide) 14 (472 mg, 1.44 mmol) and NaHSO₃ (299 mg, 2.88 mmol) in DMF (10 mL) was heated at 125° C. overnight. The mixture was poured into water (10 mL) and the resulting precipitate was collected by filtration, washed with water then triturated with MeOH to give the title compound (300 mg 17%) as a brown solid. LCMS-C (ES-API): rt 0.59 min; m/z 635.2 [M−H]⁻. ¹H NMR (400 MHz, DMSO-d₆) δ 8.15 (d, J=1.6 Hz, 2H), 7.97 (br s, 2H), 7.91 (d, J=7.8 Hz, 2H), 7.66 (dd, J=8.4, 1.7 Hz, 2H), 7.54 (t, J=7.6 Hz, 2H), 7.28 (br s, 2H), 7.14-7.12 (m, 4H), 6.63 (d, J=7.4 Hz, 2H), 4.20 (s, 4H).

Example 6—1,1′-(Ethane-1,2-diyl)bis(2-(2-(2H-tetrazol-5-yl)phenyl)-4-methoxy-1H-benzo[d]imidazole-5-carboxamide) (6)

A mixture of 4,4′-(ethane-1,2-diylbis(azanediyl))bis(3-amino-2-methoxybenzamide) 113 (50 mg, 0.13 mmol), 2-(2H-tetrazol-5-yl)benzaldehyde A8 (45 mg, 0.26 mmol) and NaHSO₃ (40 mg, 0.39 mmol) in DMF (5 mL) was heated at 120° C. under nitrogen overnight. The mixture was cooled to room temperature, poured into water (50 mL) and EtOAc (50 mL) was added. The resulting precipitate was collected by filtration and purified by prep-HPLC (Varian-940-LC; BOSTON, 10 μm, 250×2.12 mm column, eluting with a gradient of ACN in water with 0.1% HCO₂H, at a flow rate of 15 mL/min) to give the title compound (4 mg, 4%) as a white solid. LCMS-C (ES-API): rt 2.36 min; m/z 697.1 [M+H]⁺. ¹H NMR (400 MHz, DMSO-d₆) δ 7.95 (d, J=7.6 Hz, 2H), 7.69 (br s, 2H), 7.59-7.55 (m, 4H), 7.44 (br s, 2H), 7.25 (t, J=7.6 Hz, 2H), 6.76 (d, J=7.6 Hz, 2H), 6.69 (d, J=8.6 Hz, 2H), 4.28 (s, 6H), 4.08 (s, 4H).

Example 7—2,2′-(Ethane-1,2-diylbis(5-carbamoyl-4-methoxy-1H-benzo[d]imidazole-1,2-diyl))bis(4-chlorobenzoic acid) (7)

(a) Methyl 2-(bromomethyl)-4-chlorobenzoate (A9)

To a solution of methyl 4-chloro-2-methylbenzoate (2.0 g, 10.8 mmol) in 0014 (40 mL) was added NBS (2.0 g, 11.9 mmol) and AIBN (1.0 g, 5.42 mmol) and the mixture was heated at 80° C. under N₂ overnight. The mixture was poured into water, extracted with EtOAc and the combined organic extracts were washed with brine, dried over Na₂SO₄, filtered and concentrated under reduced pressure. The residue was purified by silica gel chromatography (Pet. Ether/EtOAc=100:1) to give the title compound (2.3 g, 82%) as a colorless oil. ¹H NMR (400 MHz, DMSO-d₆) δ 7.85 (d, J=2.2 Hz, 1H), 7.69-7.62 (m, 2H), 4.99 (s, 2H), 3.88 (s, 3H).

(b) Methyl 4-chloro-2-formylbenzoate (A10)

To a solution of methyl 2-(bromomethyl)-4-chlorobenzoate (A9) (2.3 g, 8.73 mmol) in THF (30 mL) was added NMO (2.0 g, 17.5 mmol) and the mixture was heated at reflux under N₂ for 16 h. Additional NMO (1.0 g, 8.73 mmol) and THF (15 mL) were added and the mixture was heated at reflux under N₂ for a further 4 h. The mixture was poured into water (40 mL), extracted with EtOAc (40 mL×3) and the combined organic extracts were washed with brine, dried over Na₂SO₄, filtered and concentrated under reduced pressure. The residue was purified by silica gel chromatography (Pet. Ether/EtOAc=200:1 to 150:1 to 120:1) to give the title compound (380 mg, 22%) as a yellow solid. LCMS-B (ES-API): rt 3.71 min, m/z 199.0, 201.0 [M+H]⁺. ¹H NMR (400 MHz, DMSO-d₆) δ 10.4 (s, 1H), 7.94-7.81 (m, 3H), 3.91 (s, 3H).

(c) Dimethyl 2,2′-(ethane-1,2-diylbis(5-carbamoyl-4-methoxy-1H-benzo[d]imidazole-1,2-diyl))bis(4-chlorobenzoate) (A11)

A mixture of methyl 4-chloro-2-formylbenzoate (A10) (0.100 g, 0.50 mmol), 4,4′-(ethane-1,2-diylbis(azanediyl))bis(3-amino-2-methoxybenzamide) (I13) (0.098 g, 0.25 mmol) and NaHSO₃ (0.052 g, 0.50 mml) in DMF (4 mL) was heated at 100° C. under N₂ for 16 h. The mixture was poured into water (10 mL), extracted with EtOAc (10 mL×3) and the combined organic extracts were washed with brine, dried over Na₂SO₄, filtered and concentrated under reduced pressure. The residue was triturated with MeOH to give the title compound (66 mg, 35%) as a white solid. LCMS-B (ES-API): rt 3.15 min, m/z 745.2, 747.2 [M+H]⁺. ¹H NMR (400 MHz, DMSO-d₆) δ 7.89 (d, J=8.5 Hz, 2H), 7.68-7.66 (m, 4H), 7.58 (dd, J=8.5, 2.1 Hz, 2H), 7.45 (d, J=1.9 Hz, 2H), 6.98 (d, J=8.6 Hz, 2H), 6.80 (br s, 2H), 4.38 (s, 6H), 4.29 (s, 4H), 3.60 (s, 6H).

(d) 2,2′-(Ethane-1,2-diylbis(5-carbamoyl-4-methoxy-1H-benzo[d]imidazole-1,2-diyl))bis(4-chlorobenzoic acid) (7)

A mixture of dimethyl 2,2′-(ethane-1,2-diylbis(5-carbamoyl-4-methoxy-1H-benzo[d]imidazole-1,2-diyl))bis(4-chlorobenzoate) (A11) (66 mg, 0.09 mmol) and NaOH (21 mg, 0.53 mmol) in EtOH/H₂O (5 mL/5 mL) was stirred at room temperature overnight. The mixture was poured into water (10 mL) and washed with EtOAc (10 mL×3). The aqueous phase was adjusted to pH 4-5 with 1 M aqueous HCl and the resulting precipitate was collected by filtration and triturated with 10:1 DCM/MeOH to give the title compound (7.4 mg, 11%) as a white solid. LCMS-B (ES-API): rt 2.75 min, m/z 717.1, 719.1 [M+H]⁺. ¹H NMR (400 MHz, DMSO-d₆) δ 13.2 (br s, 2H), 7.91 (d, J=8.5 Hz, 2H), 7.67 (d, J=2.0 Hz, 2H), 7.60 (d, J=8.6 Hz, 2H), 7.52 (dd, J=8.5, 1.8 Hz, 2H), 7.43 (d, J=2.0 Hz, 2H), 6.80 (d, J=8.6 Hz, 2H), 6.56 (br s, 2H), 4.41 (s, 6H), 4.27 (s, 4H).

Example 8—6,6′-(Ethane-1,2-diylbis(5-carbamoyl-4-methoxy-1H-benzo[d]imidazole-1,2-diyl))bis(3,4-dichlorobenzoic acid) (8)

(a) 4,5-Dichloro-2-(methoxycarbonyl)benzoic acid (A12)

A solution of 5,6-dichloroisobenzofuran-1,3-dione (5 g, 23 mmol) in methanol (50 mL) was heated to reflux for 5 h. The mixture was concentrated under reduced pressure to afford the title compound (5.7 g, 100%) as a white solid. LCMS-B (ES-API): rt 3.45 min; m/z 249.0/251.0 [M+H]⁺. ¹H NMR (400 MHz, DMSO-d₆) δ 8.00 (s, 1H), 7.96 (s, 1H), 3.81 (s, 3H).

(b) 5,6-Dichloroisobenzofuran-1(3H)-one (A13)

To a solution of 4,5-dichloro-2-(methoxycarbonyl)benzoic acid (A12) (1.0 g, 4.0 mmol) and Et₃N (1.1 mL, 8.0 mmol) in dry THF (20 mL) at 0° C. was added ClCO₂Et (654 mg, 6.0 mmol) dropwise. The mixture was stirred for 40 min, then filtered. The filtrate was diluted with dry THF (10 mL) and water (0.36 mL) was added followed by NaBH₄ (303 mg, 8.0 mmol) portion wise and the mixture was stirred until TLC analysis showed all the starting material was consumed. The mixture was filtered and the filtrate was concentrated under reduced pressure to give the title compound (812 mg, 87%) as a white solid. LCMS-B (ES-API): rt 3.61 min, m/z 202.9, 205.0 [M+H]⁺. ¹H NMR (400 MHz, DMSO-d₆) δ 8.14 (s, 1H), 8.05 (s, 1H), 5.41 (s, 2H).

(c) 3-bromo-5,6-dichloroisobenzofuran-1(3H)-one (A14)

A mixture of 5,6-dichloroisobenzofuran-1(3H)-one (A13) (580 mg, 2.86 mmol), NBS (610 mg, 3.43 mmol) and AIBN (46.9 mg, 0.286 mmol) in benzene (20 mL) was heated at reflux under N₂ overnight. The mixture was concentrated under reduced pressure and the residue was purified by silica gel chromatography (Pet. Ether/EtOAc=100:1 to 50:1) to give the title compound (545 mg, 68%) as a brown solid. ¹H NMR (400 MHz, DMSO-d₆) δ 8.19 (s, 1H), 8.11 (s, 1H), 7.99 (s, 1H).

(d) 4,5-Dichloro-2-formylbenzoic acid (A15)

A mixture of 3-bromo-5,6-dichloroisobenzofuran-1(3H)-one (A14) (547 mg, 1.95 mmol) in water (10 mL) was heated at reflux for 90 min. The water was then removed by freeze drying to give the title compound (410 mg, 96%) as a white solid. LCMS-B (ES-API): rt 3.29 min, m/z 218.9, 220.9 [M+H]⁺. 1H NMR (400 MHz, DMSO-d₆) δ 11.1 (br s, 2H), 8.09 (s, 1H), 7.96 (s, 1H).

(e) 6,6′-(Ethane-1,2-diylbis(5-carbamoyl-4-methoxy-1H-benzo[d]imidazole-1,2-diyl))bis(3,4-dichlorobenzoic acid) (8)

A mixture of 4,5-dichloro-2-formylbenzoic acid (A15) (0.100 g, 0.459 mmol), 4,4′-(ethane-1,2-diylbis(azanediyl))bis(3-amino-2-methoxybenzamide) (I13) (0.089 g, 0.23 mmol) and NaHSO₃ (0.119 g, 1.15 mmol) in DMF (3 mL) was heated at 80° C. under N₂ overnight. The mixture was concentrated by freeze drying and the residue was purified by prep. HPLC (Agilent, YMC-C18, 150×20 mm, 5 μm column, eluting with a gradient of ACN in water with 0.1% formic acid at a flow rate of 15.0 mL/min) to give the title compound (13.7 mg, 8%) as a white solid. LCMS-B (ES-API): rt 2.90 min, m/z 785.3, 787.3 [M+H]⁺. ¹H NMR (400 MHz, DMSO-d₆) δ 7.96 (s, 2H), 7.66 (s, 2H), 7.65 (d, J=8.6 Hz, 2H), 7.43 (br s, 2H), 6.94 (d, J=8.6 Hz, 2H), 6.88 (br s, 2H), 4.41 (s, 6H), 4.31 (s, 4H).

Example 9—1,1′-(Ethane-1,2-diyl)bis(2-(4-chloro-2-(2H-tetrazol-5-yl)phenyl)-4-methoxy-1H-benzo[d]imidazole-5-carboxamide) (9)

(a) 5-chloro-2-(dimethoxymethyl)benzonitrile (A16)

To a solution of 5-chloro-2-formylbenzonitrile (2.0 g, 12.1 mmol) in MeOH (60 mL) was added trimethoxymethane (2.51 g, 24.2 mmol) and TsOH (0.21 g, 1.21 mmol) and the mixture was heated at 80° C. under N₂ overnight. The solvent was removed under reduced pressure and the residue was dissolved in EtOAc, washed with a saturated aqueous NaHCO₃ solution, filtered and concentrated under reduced pressure. The residue was purified by silica gel chromatography (Pet. Ether/EtOAc=100:1 to 60:1) to give the title compound (1.7 g, 66%) as a yellow oil. LCMS-A (ES-API): rt 3.86 min, m/z 234.0, 236.0 [M+Na]⁺. ¹H NMR (400 MHz, DMSO-d₆) δ 8.01 (d, J=2.2 Hz, 1H), 7.75 (dd, J=8.4, 2.2 Hz, 1H), 7.58 (d, J=8.4 Hz, 1H), 5.50 (s, 1H), 3.26 (s, 6H).

(b) 5-(5-Chloro-2-(dimethoxymethyl)phenyl)-2H-tetrazole (A17)

To a solution of 5-chloro-2-(dimethoxymethyl)benzonitrile (A16) (1.0 g, 4.73 mmol) in NMP (20 mL) was added Et₃N.HCl (1.95 g, 14.2 mmol) and NaN₃ (1.54 g, 23.6 mmol) and the mixture was heated at 110° C. under N₂ for 3 h. Most of the NMP was removed by lyophilisation and the residue was dissolved in DCM/MeOH (10:1) and filtered. The filtrate was concentrated under reduced pressure and the residue was purified by silica gel chromatography (DCM/MeOH=50:1 to 10:1) to give the title compound (1.1 g, containing some NMP) as a yellow oil, which was used in the next step without further purification. LCMS-B (ES-API): rt 3.2 min, m/z 277.1, 279.1 [M+Na]⁺. ¹H NMR (400 MHz, DMSO-d₆) δ 7.85 (d, J=2.2 Hz, 1H), 7.68 (d, J=8.4 Hz, 1H), 7.62 (dd, J=8.4, 2.2 Hz, 1H), 6.07 (s, 1H), 3.22 (s, 6H).

(c) 4-Chloro-2-(2H-tetrazol-5-yl)benzaldehyde (A18)

A mixture of 5-(5-chloro-2-(dimethoxymethyl)phenyl)-2H-tetrazole (A17) (1.1 g) and 2 M aqueous H₂SO₄ (21.0 mL, 42.0 mmol) in acetone (30 mL) was stirred at room temperature for 3 h. The mixture was extracted with EtOAc and the combined organic extracts were washed with water and brine, dried over Na₂SO₄, filtered and concentrated under reduced pressure. The residue was purified by prep. TLC (DCM/MeOH=5:1) to give the title compound (0.66 g, 67% over two steps) as a white solid. LCMS-B (ES-API): rt 2.83 min, m/z 209.0, 211.0 [M+H]⁺. 1H NMR (400 MHz, DMSO-d₆) δ 10.7 (s, 1H), 8.08 (d, J=2.2 Hz, 1H), 7.86 (d, J=8.4 Hz, 1H), 7.58 (dd, J=8.4, 2.1 Hz, 1H).

(d) 1,1′-(Ethane-1,2-diyl)bis(2-(4-chloro-2-(2H-tetrazol-5-yl)phenyl)-4-methoxy-1H-benzo[d]imidazole-5-carboxamide) (9)

To a solution of 4-chloro-2-(2H-tetrazol-5-yl)benzaldehyde (A18) (0.100 g, 0.481 mmol) in DMF (6 mL) was added 4,4′-(ethane-1,2-diylbis(azanediyl))bis(3-amino-2-methoxybenzamide) (I13) (0.093 mg, 0.240 mmol) and NaHSO₃ (0.050 mg, 0.481 mmol) and the mixture was heated at 80° C. under N₂ overnight. The mixture concentrated under reduced pressure and the residue was purified by prep. HPLC (Varian-940-LC, YMC-C18, 5 μm, 150×20 mm column, eluting with a gradient of ACN in water with 0.1% formic acid, at a flow rate of 15 mL/min) to give the title compound (50 mg, 27%) as a white solid. LCMS-A (ES-API): rt 2.57 min, m/z 765.2, 767.2 [M+H]⁺. ¹H NMR (400 MHz, DMSO-d₆) δ 7.95 (s, 2H), 7.73 (s, 2H), 7.62 (d, J=8.5 Hz, 2H), 7.41 (s, 2H), 6.98 (d, J=7.7 Hz, 2H), 6.72 (d, J=8.4 Hz, 2H), 6.56 (d, J=8.0 Hz, 2H), 4.38 (s, 6H), 3.88 (s, 4H).

Example 10—2,2′-(Ethane-1,2-diylbis(5-carbamoyl-4-methoxy-1H-benzo[d]imidazole-1,2-diyl))bis(3-fluorobenzoic acid) (10)

(a) Ethyl 3-fluoro-2-methylbenzoate (A19)

To a solution of 3-fluoro-2-methylbenzoic acid (5.0 g, 32.5 mmol) in DMF (80 mL) was added K₂CO₃ (13.5 g, 97.5 mmol) and EtI (7.6 g, 48.8 mmol) and the mixture was stirred at room temperature under N₂ for 3 h. The mixture was diluted with water, extracted with EtOAc and the combined organic extracts were washed with brine, dried over Na₂SO₄, filtered and concentrated under reduced pressure. The residue was purified by silica gel chromatography (Pet. Ether/EtOAc=1:0 to 100:1) to give the title compound (5.4 g, 91%) as a yellow oil. LCMS-B (ES-API): rt 4.12 min, m/z 183.1 [M+H]⁺. ¹H NMR (400 MHz, DMSO-d₆) δ 7.61 (d, J=7.2 Hz, 1H), 7.43-7.30 (m, 2H), 4.30 (q, J=7.1 Hz, 2H), 2.39 (d, J=2.3 Hz, 3H), 1.31 (t, J=7.1 Hz, 3H).

(b) Ethyl 2-(bromomethyl)-3-fluorobenzoate (A20)

To a solution of ethyl 3-fluoro-2-methylbenzoate (A19) (5.4 g, 29.7 mmol) in CCl₄ (100 mL) was added NBS (6.33 g, 35.6 mmol) and AIBN (2.43 g, 14.8 mmol) and the mixture was heated at 80° C. overnight. The mixture was diluted with water, extracted with EtOAc and the combined organic extracts were washed with brine, dried over Na₂SO₄, filtered and concentrated under reduced pressure. The residue was purified by silica gel chromatography (Pet. Ether/EtOAc=1:0 to 100:1) to give the title compound (6.2 g, 86%) as a yellow oil. ¹H NMR (400 MHz, DMSO-d₆) δ 7.75-7.71 (m, 1H), 7.57-7.49 (m, 2H), 4.95 (d, J=1.7 Hz, 2H), 4.35 (q, J=7.1 Hz, 2H), 1.34 (t, J=7.1 Hz, 3H).

(c) Ethyl 3-fluoro-2-formylbenzoate (A21)

To a solution of ethyl 2-(bromomethyl)-3-fluorobenzoate (A20) (6.2 g, 23.8 mmol) in THF (20 mL) was added NMO (11.2 g, 95.4 mmol) and the mixture was heated at reflux overnight. The mixture was diluted with water, extracted with EtOAc and the combined organic extracts were washed with brine, dried over Na₂SO₄, filtered and concentrated under reduced pressure. The residue was purified by silica gel chromatography (Pet. Ether/EtOAc=1:0 to 50:1) to give the title compound (3.6 g, 77%) as a colorless oil. ¹H NMR (400 MHz, DMSO-d₆) δ 10.3 (s, 1H), 7.79-7.72 (m, 1H), 7.69-7.57 (m, 2H), 4.34 (q, J=7.1 Hz, 2H), 1.31 (t, J=7.1 Hz, 3H).

(d) Diethyl2,2′-(ethane-1,2-diylbis(5-carbamoyl-4-methoxy-1H-benzo[d]imidazole-1,2-diyl))bis(3-fluorobenzoate) (A22)

To a solution of ethyl 3-fluoro-2-formylbenzoate (A21) (0.800 g, 4.49 mmol) in DMF (15 mL) was added 4,4′-(ethane-1,2-diylbis(azanediyl))bis(3-amino-2-methoxybenzamide (I13) (0.872 g, 2.25 mmol) and NaHSO₃ (0.930 g, 8.98 mmol) and the mixture was heated at 100° C. overnight. Additional ethyl 3-fluoro-2-formylbenzoate (A21) (0.100 g, 0.51 mmol) was added and the mixture was heated at 100° C. overnight. The mixture was diluted with water, extracted with EtOAc and the combined organic extracts were washed with brine, dried over Na₂SO₄, filtered and concentrated under reduced pressure. The solid grey residue was rinsed with Pet. Ether/EtOAc=10:1 (50 mL×2) to give the title compound (1.1 g, 75%) as a grey solid. LCMS-B (ES-API): rt 3.12 min, m/z 741.4 [M+H]⁺. ¹H NMR (400 MHz, DMSO-d₆) δ 7.84 (d, J=8.6 Hz, 2H), 7.80-7.60 (m, 6H), 7.56-7.36 (m, 6H), 4.80 (br s, 2H), 4.30 (s, 6H), 4.11 (br s, 2H), 3.99-3.85 (m, 4H), 0.77 (t, J=6.4 Hz, 6H).

(e) 2,2′-(Ethane-1,2-diylbis(5-carbamoyl-4-methoxy-1H-benzo[d]imidazole-1,2-diyl))bis(3-fluorobenzoic acid) (10)

To a solution of diethyl 2,2′-(ethane-1,2-diylbis(5-carbamoyl-4-methoxy-1H-benzo[d]imidazole-1,2-diyl))bis(3-fluorobenzoate) (A22) (1.1 g, 1.49 mmol) in 5:1 EtOH/water (6 mL) was added NaOH (0.600 g, 14.9 mmol) and mixture was stirred at room temperature for 16 h. Most of the EtOH was removed under reduced pressure and the aqueous residue was adjusted to pH 5 with 1 M aqueous HCl. The resulting precipitate was collected by filtration and purified by prep. HPLC (Varian-940-LC, YMC-C18, 150×20 mm, 5 μm column, eluting with a gradient of ACN in water with 0.1% formic acid at a flow rate of 15.0 mL/min) to give the title compound (17 mg, 2%) as a white solid. LCMS-B (ES-API): rt 2.49 min, m/z 685.2 [M+H]⁺. ¹H NMR (400 MHz, DMSO-d₆) δ 13.2 (s, 2H), 7.80-7.61 (m, 8H), 7.46 (s, 2H), 7.29-7.21 (m, 4H), 4.49 (br s, 2H), 4.31 (s, 6H), 4.17 (br s, 2H).

Example 11—2,2′-(Ethane-1,2-diylbis(5-carbamoyl-4-methoxy-1H-benzo[d]imidazole-1,2-diyl))bis(5-chloro-3-fluorobenzoic acid) (11)

(a) 2-Amino-5-chloro-3-fluorobenzoic acid (A23)

A mixture of 2-amino-3-fluorobenzoic acid (5.0 g, 0.03 mol) and NCS (5.6 g, 0.04 mmol) in DMF (80 mL) was stirred at room temperature under N₂ overnight. DCM (60 mL) was added and the mixture was washed with water (2×60 mL). During the second wash, a precipitate formed, which was collected by filtration and washed with water then dried under vacuum to give the title compound (4.0 g, 71%) as a brown solid. LCMS-B (ES-API): rt 3.3 min, m/z 189.9, 191.9 [M+H]⁺. ¹H NMR (400 MHz, DMSO-d₅) δ 7.51-7.50 (m, 1H), 7.44 (dd, J=11.2, 2.5 Hz, 1H).

(b) 2-Bromo-5-chloro-3-fluorobenzoic acid (A24)

To a solution of 2-amino-5-chloro-3-fluorobenzoic acid (A23) (4.0 g, 0.021 mol) in ACN (100 mL) at 0° C. was added 40% aqueous HBr (16.9 g, 0.083 mol) dropwise over 10 min. A solution of NaNO₂ (1.6 g, 0.023 mol) in water (20 mL) was then added dropwise over 1 h and the mixture was stirred at 0° C. for 5 min. CuBr (3.6 g, 0.025 mol) was then added portion wise over 30 min and the mixture was heated at 70° C. for 1 h. Water (80 mL) was added and the resulting precipitate was collected by filtration and dried under vacuum to give the title compound (2.0 g, 41%) as a brown solid. LCMS-B (ES-API): rt 3.4 min, m/z 252.9, 254.8 [M+H]⁺. ¹H NMR (400 MHz, DMSO-d₆) δ 14.0 (br s, 1H), 7.81 (dd, J=8.6, 2.4 Hz, 1H), 7.66 (dd, J=2.4, 1.4 Hz, 1H).

(c) 5-Chloro-3-fluoro-2-formylbenzoic acid (A25)

To a solution of 2-bromo-5-chloro-3-fluorobenzoic acid (A24) (2.0 g, 0.008 mol) in anhydrous THF (50 mL) at −30° C. was added i-PrMgCl (2 M solution in THF, 16.0 mL, 0.032 mol) dropwise and the mixture was stirred for 2 h. DMF (6 mL) was then added and the mixture was allowed to warm to room temperature and stirred for 2 h. The solvent was removed under reduced pressure and the residue was diluted with 1 M aqueous HCl and extracted with EtOAc (30 mL×3). The combined organic extracts were washed with brine and concentrated under reduced pressure. The residue was purified by C₁₈ reverse phase chromatography (Biotage) eluting with a gradient of water/MeOH to give the title compound (1.1 g, 69%) as a brown solid. ¹H NMR analysis showed the lactol form of the product. LCMS-B (ES-API): rt 2.90 min, m/z 202.9, 204.9 [M+H]⁺. ¹H NMR (400 MHz, DMSO-d₆) δ 8.48 (br s, 1H), 7.91 (dd, J=8.8, 1.5 Hz, 1H), 7.80 (d, J=1.5 Hz, 1H), 6.85 (br s, 1H).

(d) 2,2′-(ethane-1,2-diylbis(5-carbamoyl-4-methoxy-1H-benzo[d]imidazole-1,2-diyl))bis(5-chloro-3-fluorobenzoic acid) (11)

A mixture of 5-chloro-3-fluoro-2-formylbenzoic acid (A25) (0.800 g, 3.95 mmol), 4,4′-(ethane-1,2-diylbis(azanediyl))bis(3-amino-2-methoxybenzamide) (I13) (0.767 g, 1.97 mmol) and NaHSO₃ (0.205 g, 1.97 mmol) in DMF (20 mL) was heated at 60° C. under N₂ overnight. Water (30 mL) was added and the resulting precipitate was collected by filtration and purified by prep. HPLC (Agilent, YMC-C18, 150×20 mm, 5 μm column, eluting with a gradient of ACN in water with 0.1% formic acid at a flow rate of 15.0 mL/min) to give the title compound (301 mg, 20%) as a grey solid. LCMS-B (ES-API): rt 2.75 min, m/z 753.3, 755.3 [M+H]⁺. ¹H NMR (400 MHz, DMSO-d₆) δ 7.76-7.63 (m, 6H), 7.47 (s, 2H), 7.42-7.24 (m, 4H), 4.68-4.43 (m, 2H), 4.31 (s, 6H), 4.25-4.05 (m, 2H).

Example 12—2,2′-(Ethane-1,2-diylbis(5-carbamoyl-4-methoxy-1H-benzo[d]imidazole-1,2-diyl)bis(4-chloro-3-fluorobenzoic acid) (12)

(a) 4-Chloro-3-fluoro-2-formylbenzoic acid (A26)

To a solution of diisopropylamine (695 mg, 6.87 mmol) in THF (20 mL) at −78° C. under N₂ was added n-BuLi (2.5 M in hexanes, 3.0 mL, 7.5 mmol) and the mixture was stirred at −78° C. for 1 h. 4-chloro-3-fluorobenzoic acid (1.0 g, 5.73 mmol) was then added and stirring was continued for a further 1 h. DMF (1.3 g, 17.2 mmol) was added and the mixture was allowed to warm to room temperature and stirred for 2 h. The reaction mixture was quenched with a saturated aqueous NH₄Cl solution and washed with EtOAc. The aqueous phase was concentrated under reduced pressure and the residue purified by silica gel chromatography (Pet. Ether to Pet. Ether/EtOAc=1:1 to DCM/MeOH=10:1) to give the title compound (1.0 g) as a white solid. LCMS analysis showed it was a ˜1:5 mixture of title compound/unreacted 4-chloro-3-fluorobenzoic acid, which was used directly in the next step without further purification. LCMS-B (ES-API): rt for title compound 2.99 min, m/z 203.0, 205.0 [M+H]⁺.

(b) 2,2′-(Ethane-1,2-diylbis(5-carbamoyl-4-methoxy-1H-benzo[d]imidazole-1,2-diyl))bis(4-chloro-3-fluorobenzoic acid) (12)

To a solution of a ˜1:5 mixture of 4-chloro-3-fluoro-2-formylbenzoic acid/4-chloro-3-fluorobenzoic acid (A26) (104 mg) and 4,4′-(ethane-1,2-diylbis(azanediyl))bis(3-amino-2-methoxybenzamide) (I13) (100 mg, 0.26 mmol) in DMF (8 mL) was added NaHSO₃ (107 mg, 1.03 mmol) and the mixture was heated at 120° C. under N₂ overnight. The mixture was purified by prep. HPLC (Agilent, YMC-C18, 150×20 mm, 5 μm column, eluting with a gradient of ACN in water with 0.1% formic acid at a flow rate of 15.0 mL/min) to give the title compound (5 mg) as a white solid. LCMS-B (ES-API): rt 2.66 min, m/z 753.1, 755.1 [M+H]⁺. ¹H NMR (400 MHz, DMSO-d₆) δ 7.77 (d, J=8.6 Hz, 2H), 7.68 (d, J=3.0 Hz, 2H), 7.63-7.59 (m, 4H), 7.42 (br s, 2H), 7.03 (d, J=8.5 Hz, 2H), 4.50-4.47 (m, 2H), 4.37 (s, 6H), 4.24-4.21 (m, 2H).

Example 13—Dimethyl 6,6′-(ethane-1,2-diylbis(5-carbamoyl-4-methoxy-1H-benzo[d]imidazole-1,2-diyl))bis(3-bromobenzoate) (13) and 6,6′-(Ethane-1,2-diylbis(5-carbamoyl-4-methoxy-1H-benzo[d]imidazole-1,2-diyl))bis(3-bromobenzoic acid) (14)

(a) Methyl 5-bromo-2-(bromomethyl)benzoate (A27)

A mixture of methyl 5-bromo-2-methylbenzoate (1.0 g, 4.4 mmol), NBS (0.56 g, 4.4 mmol) and AIBN (0.36 g, 2.2 mmol) in CCl₄ (20 mL) was heated at 80° C. under N₂ for 2 h. Additional NBS (56.3 mg, 0.44 mmol) was added and heating was continued at 80° C. for 1 h. Additional NBS (56.3 mg, 0.44 mmol) was added heating was continued at 80° C. for 1 h. Water (30 mL) was added and the mixture was extracted with EtOAc (30 mL×3). The combined organic extracts were washed with brine, dried over Na₂SO₄, filtered and concentrated under reduced pressure. The residue was purified by silica gel chromatography (Pet. Ether/EtOAc=120:1 to 80:1) to give the title compound (1.01 g, 75%) as a yellow solid. ¹H NMR (400 MHz, DMSO-d₆) δ 7.98 (d, J=2.2 Hz, 1H), 7.80 (dd, J=8.3, 2.2 Hz, 1H), 7.56 (d, J=8.3 Hz, 1H), 4.98 (s, 2H), 3.88 (s, 3H).

(b) Methyl 5-bromo-2-formylbenzoate (A28)

A solution of methyl 5-bromo-2-(bromomethyl)benzoate (A27) (1.01 g, 3.3 mmol) and NMO (1.55 g, 13.2 mmol) in THF (20 mL) was heated at reflux under N₂ overnight. Water (20 mL) was added and the mixture was extracted with EtOAc (20 mL×3). The combined organic extracts were washed with brine, dried over Na₂SO₄, filtered and concentrated under reduced pressure. The residue was purified by silica gel chromatography (Pet. Ether/EtOAc=100:1 to 70:1) to give the title compound (217 mg, 26%) as a yellow solid. LCMS-B (ES-API): rt 3.7 min, m/z 243.0, 245.0 [M+H]⁺. ¹H NMR (400 MHz, DMSO-d₆) δ 10.3 (s, 1H), 8.06 (d, J=2.0 Hz, 1H), 8.00 (dd, J=8.5, 2.2 Hz, 1H), 7.80 (d, J=8.3 Hz, 1H), 3.91 (s, 3H).

(c) Dimethyl 6,6′-(ethane-1,2-diylbis(5-carbamoyl-4-methoxy-1H-benzo[d]imidazole-1,2-diyl))bis(3-bromobenzoate) (13)

A mixture of methyl 5-bromo-2-formylbenzoate (A28) (100 mg, 0.41 mmol), 4,4′-(ethane-1,2-diylbis(azanediyl))bis(3-amino-2-methoxybenzamide) (I13) (81.6 mg, 0.21 mmol) and NaHSO₃ (21.9 mg, 0.21 mmol) in DMF (5 mL) was heated at 120° C. under N₂ overnight. Water (20 mL) was added and resulting precipitate was collected by filtration and dried under vacuum to give the title compound (66 mg, 38%) as a black solid. LCMS-B (ES-API): rt 3.17 min, m/z 833.1, 835.1 [M+H]⁺. ¹H NMR (400 MHz, DMSO-d₆) δ 7.94 (d, J=2.1 Hz, 2H), 7.72-7.69 (m, 4H), 7.46-7.41 (m, 4H), 7.11 (d, J=8.7 Hz, 2H), 6.77 (d, J=8.0 Hz, 2H), 4.36 (s, 6H), 4.25 (s, 4H), 3.61 (s, 6H).

(d) 6,6′-(Ethane-1,2-diylbis(5-carbamoyl-4-methoxy-1H-benzo[d]imidazole-1,2-diyl))bis(3-bromobenzoic acid) (14)

To a solution of dimethyl 6,6′-(ethane-1,2-diylbis(5-carbamoyl-4-methoxy-1H-benzo[d]imidazole-1,2-diyl))bis(3-bromobenzoate) (13) (60 mg, 0.07 mmol) in 9:1 EtOH/water (5.5 mL) was added NaOH (28 mg, 0.7 mmol) and the mixture was stirred at room temperature overnight. Most of the EtOH was removed under reduced pressure and the aqueous residue was adjusted to pH 5 with 2 M aqueous HCl. The resulting precipitate was collected by filtration and dried under vacuum to give the title compound (35 mg, 62%) as a black solid. LCMS-B (ES-API): rt 2.66 min, m/z 805.1, 807.0 [M+H]⁺. ¹H NMR (400 MHz, DMSO-d₆) δ 7.95 (d, J=2.0 Hz, 2H), 7.72 (br s, 2H), 7.65 (d, J=8.6 Hz, 2H), 7.45 (br s, 2H), 7.33 (dd, J=8.0, 1.6 Hz, 2H), 6.98 (d, J=8.6 Hz, 2H), 6.60 (d, J=8.2 Hz, 2H), 4.37 (s, 6H), 4.25 (s, 4H).

Example 15—6,6′-(Ethane-1,2-diylbis(5-carbamoyl-4-methoxy-1H-benzo[d]imidazole-1,2-diyl))bis(3,4-difluorobenzoic acid) (15)

(a) 4,5-Difluoro-2-formylbenzoic acid (A29)

To a solution of 2-bromo-4,5-difluorobenzoic acid (500 mg, 2.11 mmol) in THF (5 mL) at −78° C. under N₂ was added n-BuLi (2.5 M solution in hexanes, 1.70 mL, 4.22 mmol) dropwise and the mixture was stirred at −78° C. for 45 min. DMF (0.20 mL, 2.53 mmol) was then added and stirring was continued at −78° C. for 2 h. The reaction was quenched with a saturated aqueous NH₄C₁ solution and the mixture was partitioned between EtOAc and water. The layers were separated and the aqueous phase was freeze dried. The residue was purified by prep. TLC (DCM/MeOH=10:1) to give the title compound (16 mg, 4%) as a white solid. LCMS-B (ES-API): rt 2.50 min; m/z 187.0 [M+H]⁺.

(b) 6,6′-(Ethane-1,2-diylbis(5-carbamoyl-4-methoxy-1H-benzo[d]imidazole-1,2-diyl))bis(3,4-difluorobenzoic acid) (15)

A mixture of 4,5-difluoro-2-formylbenzoic acid (A29) (16 mg, 0.086 mmol), 4,4′-(ethane-1,2-diylbis(azanediyl))bis(3-amino-2-methoxybenzamide) (I13) (23 mg, 0.07 mmol) and NaHSO₃ (14 mg, 0.13 mml) in DMF (1 mL) was heated at 120° C. overnight. The mixture was partitioned between EtOAc and water, the layers were separated the aqueous layer was freeze dried. The residue was purified by prep. HPLC (Agilent, YMC-C18, 150×20 mm, 5 μm column, eluting with a gradient of ACN in water with 0.1% formic acid at a flow rate of 15.0 mL/min) to give the title compound (10 mg, 32%) as a white solid. LCMS-B (ES-API): rt 2.54 min; m/z 721.2 [M+H]⁺. 1H NMR (400 MHz, DMSO-d₆) δ 7.81 (dd, J=8.3, 4.9 Hz, 2H), 7.68 (d, J=2.0 Hz, 2H), 7.60 (d, J=7.9 Hz, 2H), 7.49-7.38 (m, 4H), 7.08 (d, J=7.7 Hz, 2H), 4.48-4.46 (m, 2H), 4.34 (s, 6H), 4.23-4.22 (m, 2H).

Example 16—2,2′-(Ethane-1,2-diylbis(5-carbamoyl-4-methoxy-1H-benzo[d]imidazole-1,2-diyl))bis(3,5-difluorobenzoic acid) (16)

(a) 3,5-Difluoro-2-formylbenzoic acid (A30)

To a solution of 2-bromo-3,5-difluorobenzoic acid (500 mg, 2.11 mmol) in anhydrous THF (10 mL) at −30° C. was added i-PrMgCl (2 M solution in THF, 2.2 mL, 4.4 mmol) and the mixture was stirred at −30° C. for 3 h. DMF (1.5 mL) was added and the mixture was allowed to warm to room temperature and stirred for 2 h. The mixture was concentrated under reduced pressure and the residue was diluted with 0.5 M aqueous HCl, extracted with DCM and the combined organic extracts were washed with brine, dried over Na₂SO₄, filtered and concentrated under reduced pressure. The residue was purified by silica gel chromatography (Pet. Ether/EtOAc=100:0 to 100:1) to give the title compound (130 mg, 33%) as a yellow solid. ¹H NMR analysis showed the lactol form of the product. LCMS-B (ES-API): rt 2.69 min, m/z 186.9 [M+H]⁺. ¹H NMR (400 MHz, DMSO-d₆) δ 8.44 (br s, 1H), 7.75 (td, J=9.3, 2.1 Hz, 1H), 7.62 (dd, J=7.0, 2.1 Hz, 1H), 6.82 (br s, 1H).

(b) 2,2′-(Ethane-1,2-diylbis(5-carbamoyl-4-methoxy-1H-benzo[d]imidazole-1,2-diyl))bis(3,5-difluorobenzoic acid) (16)

A mixture of 3,5-difluoro-2-formylbenzoic acid (A30) (130 mg, 0.70 mmol), 4,4′-(ethane-1,2-diylbis(azanediyl))bis(3-amino-2-methoxybenzamide) (I13) (136 mg, 0.35 mmol) and NaHSO₃ (73 mg, 0.70 mmol) in DMF (8 mL) was heated at 120° C. under N₂ for 16 h. The mixture was concentrated under reduced pressure until a precipitate formed, which was collected by filtration and purified by prep. HPLC (Agilent, YMC-C18, 150×20 mm, 5 μm column, eluting with a gradient of ACN in water with 0.1% formic acid at a flow rate of 15.0 mL/min) to give the title compound (2.5 mg, 1%) as a white solid. LCMS-B (ES-API): rt 2.55 min, m/z 721.3 [M+H]⁺. ¹H NMR (400 MHz, DMSO-d₆) δ 7.76-7.62 (m, 4H), 7.60-7.50 (m, 2H), 7.47 (d, J=2.8 Hz, 2H), 7.30-7.06 (m, 2H), 7.26 (d, J=8.6 Hz, 2H), 4.63-4.42 (m, 2H), 4.32 (s, 6H), 4.24-4.08 (m, 2H). ¹H NMR (400 MHz, 60° C., DMSO-d₆) δ 7.75 (d, J=8.6 Hz, 2H), 7.66-7.51 (m, 4H), 7.38-7.16 (m, 4H), 7.20 (d, J=8.6 Hz, 2H), 4.43-4.24 (m, 10H).

Example 17—1,1′-(Ethane-1,2-diyl)bis(2-(4-chloro-2-(5-oxo-4,5-dihydro-1,3,4-oxadiazol-2-yl)phenyl)-4-methoxy-1H-benzo[d]imidazole-5-carboxamide) (17)

(a) Di-tert-butyl 2,2′-(2,2′-(ethane-1,2-diylbis(5-carbamoyl-4-methoxy-1H-benzo[d]imidazole-1,2-diyl))bis(5-chlorobenzoyl))bis(hydrazine-1-carboxylate) (A31) A mixture of 6,6′-(ethane-1,2-diylbis(5-carbamoyl-4-methoxy-1H-benzo[d]imidazole-1,2-diyl))bis(3-chlorobenzoic acid) (1) (0.300 g, 0.42 mmol), tert-butyl hydrazinecarboxylate (0.332 g, 2.52 mmol), EDCl.HCl (0.240 g, 1.26 mmol), HOBt (0.169 g, 1.26 mmol) and DIPEA (0.324 g, 2.52 mmol) in DMF (12 mL) was stirred at room temperature under N₂ for 16 h. The mixture poured into water (30 mL) and extracted with EtOAc (30 mL×3). The combined organic extracts were washed water (30×2 mL), brine (30 mL), dried over Na₂SO₄, filtered and concentrated under reduced pressure to give the title compound (410 mg, >100%) as a yellow oil, which was used directly in the next step without further purification.

(b) 1,1′-(Ethane-1,2-diyl)bis(2-(4-chloro-2-(hydrazinecarbonyl)phenyl)-4-methoxy-1H-benzo[d]imidazole-5-carboxamide) dihydrochloride (A32)

A mixture of 2,2′-(2,2′-(ethane-1,2-diylbis(5-carbamoyl-4-methoxy-1H-benzo[d]imidazole-1,2-diyl))bis(5-chlorobenzoyl))bis(hydrazine-1-carboxylate) (A31) (410 mg, assumed quant. yield from previous step, 0.42 mmol) and a 3 M HCl in dioxane solution (6 mL) was stirred at room temperature for 4 h. The mixture was concentrated under reduced pressure to give the title compound (400 mg, >100%) as a grey solid, which was used directly in the next step without further purification. LCMS-B (ES-API): rt 1.67 min, m/z 745.3, 747.3 [M+H]⁺.

(c) 1,1′-(Ethane-1,2-diyl)bis(2-(4-chloro-2-(5-oxo-4,5-dihydro-1,3,4-oxadiazol-2-yl)phenyl)-4-methoxy-1H-benzo[d]imidazole-5-carboxamide) (17)

A mixture of 1,1′-(ethane-1,2-diyl)bis(2-(4-chloro-2-(hydrazinecarbonyl)phenyl)-4-methoxy-1H-benzo[d]imidazole-5-carboxamide) dihydrochloride (A32) (90 mg, assumed quant. yield from previous step, 0.095 mmol), CDI (59 mg, 0.36 mmol) and Et₃N (37 mg, 0.36 mmol) in DMF (4 mL) was heated at 40° C. under N₂ for 16 h. The mixture poured into water (20 mL), extracted with EtOAc (15 mL×3) and the combined organic extracts were washed with water (15×2 mL), brine (15 mL), dried over Na₂SO₄, filtered and concentrated under reduced pressure. The residue was purified by prep. HPLC (Agilent, BOSTON, 250×2.12 mm, 10 μm column, eluting with a gradient of ACN in water with 0.1% formic at a flow rate of 20.0 mL/min) to give the title compound (5 mg, 7%) as a grey solid. LCMS-B (ES-API): rt 2.75 min, m/z 797.4, 799.4 [M+H]⁺. ¹H NMR (400 MHz, DMSO-d₆) δ 12.6 (br s, 2H), 7.71 (br s, 2H), 7.66 (d, J=1.8 Hz, 2H), 7.60 (d, J=8.6 Hz, 2H), 7.46 (br s, 2H), 7.05 (d, J=6.8 Hz, 2H), 6.96 (d, J=8.6 Hz, 2H), 6.64 (d, J=8.3 Hz, 2H), 4.38 (s, 6H), 4.21 (s, 4H).

Example 18—2,2′-(Ethane-1,2-diylbis(5-carbamoyl-7-fluoro-4-methoxy-1H-benzo[d]imidazole-1,2-diyl)bis(3,5-difluorobenzoic acid) (18)

(a) 4,5-Difluoro-2-methoxybenzoic acid (A33)

Prepared from 2,4,5-trifluorobenzoic acid according to the procedure described in Bioorg. Med. Chem. Lett. 2016, 26, 2455. LCMS-B (ES-API): rt 2.98 min, m/z 189.0 [M+H]⁺. ¹H NMR (400 MHz, CDCl₃) δ 7.99 (dd, J=10.5, 9.1 Hz, 1H), 6.90 (dd, J=11.2, 6.0 Hz, 1H), 4.05 (s, 3H).

(b) 4,5-Difluoro-2-methoxybenzamide (A34)

To a solution of 4,5-difluoro-2-methoxybenzoic acid (A33) (16.5 g, 85 mmol) in dry DCM (300 mL) at 0° C. was added DMF (10 drops) and (COCl)₂ (15 mL, 17 mmol) and the mixture was allowed to warm to room temperature and stirred for 3 h. The solvent was removed under reduced pressure and the residue was dissolved in dry DCM (30 mL) and cooled to 0° C. Conc. aqueous NH₄OH (26-28% aqueous solution, 70 mL) was added and the mixture was stirred at room temperature overnight. Most of the DCM was removed under reduced pressure and the resulting precipitate was collected by filtration and dried at 50° C. overnight to give the title compound (15.5 g, 97%) as a white solid. LCMS-B (ES-API): rt 2.76 min, m/z 188.0. [M+H]⁺. ¹H NMR (400 MHz, DMSO-d₆) δ 7.76 (dd, J=11.5, 9.6 Hz, 1H), 7.70 (br s, 1H), 7.66 (br s, 1H), 7.32 (dd, J=12.7, 6.5 Hz, 1H), 3.89 (s, 3H).

(c) 4,5-Difluoro-2-methoxy-3-nitrobenzamide (A35)

To a solution of 4,5-difluoro-2-methoxybenzamide (A34) (12.0 g, 64.2 mmol) in conc. H₂SO₄ (150 mL) was added a mixture of KNO₃ (9.74 g, 96.3 mmol) in conc. H₂SO₄ (5 mL) dropwise over 30 min, maintaining the reaction temperature at 0° C. Following complete addition, TLC analysis showed the starting material was consumed. The mixture was diluted with water, extracted with EtOAc and the combined organic extracts were washed with brine, dried over Na₂SO₄, filtered and concentrated under reduced pressure. The residue was purified by silica gel chromatography (DCM/MeOH=1:0 to 200:1 to 100:1) to give the title compound (4.5 g, 30%) as a white solid. LCMS-B (ES-API): rt 3.12 min, m/z 232.9 [M+H]⁺. ¹H NMR (400 MHz, DMSO-d₆) δ 7.99 (br s, 1H), 7.96 (dd, J=10.4, 8.8 Hz, 1H), 7.92 (br s, 1H), 3.89 (s, 3H).

(d) 4,4′-(Ethane-1,2-diylbis(azanediyl))bis(5-fluoro-2-methoxy-3-nitrobenzamide) (A36)

To a solution of 4,5-difluoro-2-methoxy-3-nitrobenzamide (A35) (2.5 g, 10.8 mmol) in DMF (13 mL) was added K₂CO₃ (4.48 g, 32.4 mmol) and ethylenediamine (324 mg, 5.4 mmol) and the mixture was stirred at room temperature overnight. The mixture was carefully poured into ice water with stirring and the resulting precipitate was collected by filtration, washed with water (10 mL) and dried at 50° C. overnight to give the title compound (1.91 g, 73%) as a yellow solid. LCMS-B (ES-API): rt 3.07 min, m/z 485.1 [M+H]⁺. ¹H NMR (400 MHz, DMSO-d₆) δ 7.56 (br s, 2H), 7.54 (br s, 2H), 7.47 (d, J=13.9 Hz, 2H), 6.27-6.20 (m, 2H), 3.75 (s, 6H), 3.34-3.33 (m, 4H).

(e) 2,2′-(Ethane-1,2-diylbis(5-carbamoyl-7-fluoro-4-methoxy-1H-benzo[d]imidazole-1,2-diyl))bis(3,5-difluorobenzoic acid) (18)

To a solution of 4,4′-(ethane-1,2-diylbis(azanediyl))bis(5-fluoro-2-methoxy-3-nitrobenzamide) (A36) (0.200 g, 0.41 mmol) in 1:1 DMSO/MeOH (8 mL) was added 3,5-difluoro-2-formylbenzoic acid (A30) (0.152 g, 0.82 mmol) and Na₂S₂O₄ (0.214 g, 1.23 mmol). The flask was fitted with a condenser and the mixture was heated at 120° C. for 6 h. The mixture was concentrated under reduced pressure to remove most of the solvent, and the residue was diluted with water (20 mL) and adjusted to pH 5 with 1 M aqueous HCl. The resulting precipitate was collected by filtration, dried under vacuum at 50° C. overnight then purified by prep. HPLC (Agilent, YMC-C18, 150×20 mm, 5 μm column, eluting with a gradient of ACN in water with 0.1% TFA at a flow rate of 15.0 mL/min) to give the title compound (20.7 mg, 7%) as a white solid. LCMS-B (ES-API): rt 2.71 min, m/z 757.3 [M+H]⁺. ¹H NMR (400 MHz, DMSO-d₆) δ 7.78 (s, 2H), 7.71-7.61 (m, 4H), 7.40-7.30 (m, 2H), 7.15 (br s, 2H), 4.50-4.36 (m, 4H), 4.24 (s, 6H).

Example 19—1,1′-(Ethane-1,2-diyl)bis(2-(5-chloro-2-(2H-tetrazol-5-yl)phenyl)-4-methoxy-1H-benzo[d]imidazole-5-carboxamide) (42)

(a) 4-Chloro-2-(dibromomethyl)benzonitrile (A37)

A solution of 4-chloro-2-methylbenzonitrile (10.0 g, 66.0 mmol), NBS (35.2 g, 198 mmol) and BPO (1.60 g, 6.60 mmol) in 0014 (200 mL) was heated at reflux under N₂ for 24 h. The mixture was concentrated under reduced pressure and the residue was purified by silica gel chromatography (Pet. ether/EtOAc=500:1 to 200:1) to give the title compound (13.0 g, 64%) as a white solid. ¹H NMR (400 MHz, DMSO-d₆) δ 8.02 (d, J=8.4 Hz, 1H), 7.91 (d, J=2.2 Hz, 1H), 7.70 (dd, J=8.3, 2.1 Hz, 1H), 7.40 (s, 1H).

(b) 4-Chloro-2-formylbenzonitrile (A38)

To a solution of 4-chloro-2-(dibromomethyl)benzonitrile A37 (10.0 g, 32.3 mmol) in acetonitrile (75 mL) was added a solution of AgNO₃ (16.5 g, 97.0 mmol) in water (17 mL) and the mixture was heated at reflux for 20 min then allowed to cool to room temperature. The precipitate was removed by filtration and washed with DCM (3×200 mL). The combined filtrates were washed with water (100 mL), dried over Na₂SO₄, filtered and concentrated under reduced pressure. The residue was purified by silica gel chromatography (Pet. ether/EtOAc=100:1 to 20:1) to give the title compound (5.30 g, 99%) as a yellow solid. LCMS-B (ES-API): rt 3.35 min, m/z 166.0, 168.0 [M+H]⁺.

(c) 4-Chloro-2-(dimethoxymethyl)benzonitrile (A39)

To a solution of 4-chloro-2-formylbenzonitrile A38 (5.00 g, 30.2 mmol) in MeOH (150 mL) was added trimethoxymethane (6.41 g, 60.4 mmol) and TsOH (520 mg, 3.02 mmol) and the mixture was heated at reflux for 16 h. The mixture was concentrated under reduced pressure and the residue was diluted with a saturated aqueous NaHCO₃ solution and extracted with EtOAc. The organic extract was dried over Na₂SO₄, filtered and concentrated under reduced pressure. The residue was purified by silica gel chromatography (Pet. ether/EtOAc=100:1 to 70:1) to give the title compound (5.30 g, 83%) as a yellow oil. LCMS-B (ES-API): rt 3.83 min, m/z 212.1, 214.1 [M+H]⁺.

(d) 5-(4-Chloro-2-(dimethoxymethyl)phenyl)-2H-tetrazole (A40)

A mixture of 4-chloro-2-(dimethoxymethyl)benzonitrile (3.00 g, 14.2 mmol), Et₃N HCl (2.73 g, 19.9 mmol) and NaN₃ (2.76 g, 42.5 mmol) in DMF (40 mL) was heated at 110° C. under N₂ for 16 h. The mixture was poured into water (200 mL) and washed with EtOAc (100 mL×3). The aqueous layer was lyophilized to dryness and the residue was purified by silica gel chromatography (DCM/MeOH=100:1 to 20:1) to give the title compound (2.20 g, 61%) as a yellow oil. LCMS-B (ES-API): rt 3.27 min, m/z 253.0, 255.0 [M−H]⁻.

(e) 5-Chloro-2-(2H-tetrazol-5-yl)benzaldehyde (A41)

To a solution of 5-(4-chloro-2-(dimethoxymethyl)phenyl)-2H-tetrazole A40 (900 mg, 3.53 mmol) in acetone (36 mL) was added a 2 M aqueous H₂SO₄ solution (36 mL, 70.6 mmol) and the mixture was stirred at room temperature for 16 h. The mixture was extracted with EtOAc and the combined organic layers were washed with water, brine, dried over Na₂SO₄, filtered and concentrated under reduced pressure to give the title compound (365 mg, 50%) as a white solid. LCMS-B (ES-API): rt 2.94 min, m/z 209.0, 211.0 [M+H]⁺. ¹H NMR (400 MHz, DMSO-d₆) δ 10.3 (s, 1H), 7.98 (d, J=2.0 Hz, 1H), 7.97-7.96 (m, 2H).

(f) 1,1′-(Ethane-1,2-diyl)bis(2-(5-chloro-2-(2H-tetrazol-5-yl)phenyl)-4-methoxy-1H-benzo[d]imidazole-5-carboxamide) (42)

A solution of 4,4′-(ethane-1,2-diylbis(azanediyl))bis(3-amino-2-methoxybenzamide) I13 (200 mg, 0.51 mmol), 5-chloro-2-(2H-tetrazol-5-yl)benzaldehyde A41 (236 mg, 0.58 mmol) and Oxone® (380 mg, 0.61 mmol) in DMF (6 mL) was stirred at room temperature under N₂ for 2 h. The mixture was poured into water (30 mL) and the resulting precipitate was collected by filtration and recrystallized from methanol to give the title compound (55 mg, 14%) as a grey solid. LCMS-B (ES-API): rt 2.76 min, m/z 765.0, 767.0 [M+H]⁺. ¹H NMR (400 MHz, DMSO-d₆) δ 7.91 (d, J=8.4 Hz, 2H), 7.67-7.65 (m, 4H), 7.61 (d, J=8.1 Hz, 2H), 7.45 (br s, 2H), 6.93-6.91 (m, 4H), 4.29 (s, 6H), 4.17 (s, 4H).

Example 20—1,1′-(Ethane-1,2-diyl)bis(2-(2,4-difluoro-6-(2H-tetrazol-5-yl)phenyl)-4-methoxy-1H-benzo[d]imidazole-5-carboxamide) (43)

(a) 2-Bromo-3,5-difluorobenzonitrile (A42)

To a solution of 3,5-difluorobenzonitrile (5.00 g, 35.9 mmol), 1,3-dibromo-5,5-dimethylimidazolidine-2,4-dione (10.3 g, 36.0 mmol) and ACN (5.90 g, 143.8 mmol) in DCM (75 mL) at 0° C. was added conc. H₂SO₄ (14.1 g, 143.8 mmol) dropwise over 30 min. The mixture was allowed to warm to room temperature then heated at 50° C. for 4 h. The mixture was partitioned between a 1 M aqueous sodium sulfite solution (120 mL) and DCM (70 mL), the layers were separated and the organic phase washed with a 1 M aqueous sodium sulfite solution (120 mL), water (70 mL), dried over Na₂SO₄, filtered and concentrated under reduced pressure. The residue was purified by silica gel chromatography (Pet. ether/EtOAc=200:1 to 100:1) to give the title compound (7.2 g, 92%) as a white solid. ¹H NMR (400 MHz, DMSO-d₆) δ 7.98-7.94 (m, 1H), 7.92 (dd, J=8.9, 2.8 Hz, 1H).

(b) 3,5-Difluoro-2-vinylbenzonitrile (A43)

To a solution of 2-bromo-3,5-difluorobenzonitrile A42 (5.00 g, 22.9 mmol) and tributyl(vinyl)stannane (21.8 g, 68.8 mmol) in toluene (100 mL) was added Pd(PPh₃)₄ (2.65 g, 2.29 mmol) and the mixture was heated at reflux under N₂ for 16 h. The mixture was poured into water (400 mL), extracted with EtOAc (200 mL×3) and the combined organic extracts were washed with water (100×2 mL), brine (100 mL), dried over Na₂SO₄, filtered and concentrated under reduced pressure. The residue was purified by silica gel chromatography (Pet. ether/EtOAc=500:1 to 200:1) to give the title compound (2.00 g, 54%) as a yellow solid. ¹H NMR (400 MHz, DMSO-d₆) δ 7.86-7.72 (m, 2H), 6.77 (dd, J=17.8, 11.8 Hz, 1H), 6.05 (d, J=17.8 Hz, 1H), 5.82 (d, J=11.7 Hz, 1H).

(c) 5-(3,5-Difluoro-2-vinylphenyl)-1H-tetrazole (A44)

A solution of 3,5-difluoro-2-vinylbenzonitrile A43 (2.00 g, 12.1 mmol), Et₃N.HCl (2.34 g, 18.8 mmol) and NaN₃ (2.36 g, 36.3 mmol) in DMF (40 mL) was heated at 110° C. under N₂ for 16 h. The mixture was poured into water (400 mL) and washed with EtOAc (250 mL×3). The aqueous phase was lyophilized to dryness and the residue was purified by silica gel chromatography (DCM/MeOH=100:1 to 20:1) to give the title compound (500 mg, 20%) as a brown solid. LCMS-B (ES-API): rt 3.23 min, m/z 209.0 [M+H]⁺.

(d) 2,4-Difluoro-6-(1H-tetrazol-5-yl)benzaldehyde (A45)

A solution of 5-(3,5-difluoro-2-vinylphenyl)-1H-tetrazole A44 (500 mg, 2.40 mmol) in DCM/MeOH (6 mL/6 mL) was cooled to −70° C. and bubbled with O₃ gas until a blue color persisted. Dimethyl sulfide (746 mg, 12.0 mmol) was then added and the mixture was allowed to warm to room temperature and stirred for 1 h. The mixture was concentrated under reduced pressure and the residue was purified by silica gel chromatography (DCM/MeOH=100:1 to 50:1) to give the title compound (360 mg, 71%) as a yellow solid. LCMS-B (ES-API): rt 2.34 min, m/z 211.0 [M+H]⁺. ¹H NMR (400 MHz, DMSO-d₆) δ 10.1 (s, 1H), 7.78-7.70 (m, 1H), 7.67 (d, J=8.9 Hz, 1H).

(e) 1,1′-(Ethane-1,2-diyl)bis(2-(2,4-difluoro-6-(2H-tetrazol-5-yl)phenyl)-4-methoxy-1H-benzo[d]imidazole-5-carboxamide) (43)

A solution of 4,4′-(ethane-1,2-diylbis(azanediyl))bis(3-amino-2-methoxybenzamide) 113 (160 mg, 0.41 mmol), 2,4-difluoro-6-(1H-tetrazol-5-yl)benzaldehyde A45 (190 mg, 0.90 mmol) and Oxone® (304 mg, 0.49 mmol) in DMF (8 mL) was stirred at room temperature under N₂ for 1 h. The mixture was poured into water (50 mL) and the resulting precipitate collected by filtration. Purification by prep-HPLC (Agilent, BOSTON 250×2.12 mm, 10 μm column, eluting with a gradient of ACN in water with 0.1% HCO₂H at a flow rate of 20.0 mL/min) gave the title compound (5 mg, 2%) as a grey solid. LCMS-B (ES-API): rt 2.57 min, m/z 769.4 [M+H]⁺. ¹H NMR (400 MHz, DMSO-d₆) δ 7.73-7.66 (m, 4H), 7.59 (d, J=8.4 Hz, 2H), 7.48-7.40 (m, 4H), 7.06-6.97 (m, 2H), 4.75-4.63 (m, 2H), 4.36-4.23 (m, 2H), 4.16 (s, 6H).

Example 21—1,1″-(Ethane-1,2-diyl)bis(2-(2,4-difluoro-6-(5-oxo-4,5-dihydro-1,3,4-oxadiazol-2-yl)phenyl)-4-methoxy-1H-benzo[d]imidazole-5-carboxamide) (44)

(a) Di-tert-butyl 2,2′-(2,2′-(ethane-1,2-diylbis(5-carbamoyl-4-methoxy-1H-benzo[d]imidazole-1,2-diyl))bis(3,5-difluorobenzoyl))bis(hydrazine-1-carboxylate) (A46)

To a solution of 2,2′-(ethane-1,2-diylbis(5-carbamoyl-4-methoxy-1H-benzo[d]imidazole-1,2-diyl))bis(3,5-difluorobenzoic acid) 16 (500 mg, 0.7 mmol) in THF (2.5 mL) was added T3P® (50% w/w in EtOAc, 2.23 g, 3.5 mmol), tert-butyl carbazate (370 mg, 2.8 mmol) and Et₃N (1.4 g, 14.0 mmol) and the mixture was stirred at room temperature overnight. The solvent was removed under reduced pressure and LCMS analysis showed incomplete reaction. The residue was dissolved in THF (2.5 mL) and T3P® (50% w/w in EtOAc, 2.23 g, 3.5 mmol), tert-butyl carbazate (370 mg, 2.8 mmol) and Et₃N (1.4 g, 14.0 mmol) were added and the mixture was heated at 80° C. for 7 h. The solvent was removed under reduced pressure and LCMS analysis showed incomplete reaction. The residue was dissolved in THF (2.5 mL) and T3P® (50% w/w in EtOAc, 2.23 g, 3.5 mmol), tert-butyl carbazate (370 mg, 2.8 mmol) and Et₃N (1.4 g, 14.0 mmol) were added and the mixture was stirred at room temperature overnight. The solvent was removed under reduced pressure and LCMS analysis showed incomplete reaction. The residue was dissolved in THF (2.5 mL) and T3P® (50% w/w in EtOAc, 2.23 g, 3.5 mmol), tert-butyl carbazate (370 mg, 2.8 mmol) and Et₃N (1.4 g, 14.0 mmol) were added and the mixture was heated at 50° C. for 4 h. The solvent was removed under pressure and the residue was washed with diethyl ether (×3) to give the title compound (290 mg, 44%) as a light brown solid. LCMS-C (ES-API): rt 3.61 min; m/z 949.4 [M+H]⁺.

(b) 1,1′-(Ethane-1,2-diyl)bis(2-(2,4-difluoro-6-(hydrazinecarbonyl)phenyl)-4-methoxy-1H-benzo[d]imidazole-5-carboxamide) hydrochloride (A47)

To a solution of di-tert-butyl 2,2′-(2,2′-(ethane-1,2-diylbis(5-carbamoyl-4-methoxy-1H-benzo [d]imidazole-1,2-diyl))bis(3,5-difluorobenzoyl))bis(hydrazine-1-carboxylate) A46 (290 mg, 0.3 mmol) in dioxane (1 mL) was added a 4 M solution of HCl in dioxane (20 mL) dropwise and the mixture was stirred at room temperature for 2 h. The mixture was concentrated under reduced pressure and the residue was washed with diethyl ether, petroleum ether and DCM to give the title compound (290 mg) as a yellow solid, which was used directly in the next step without further purification. LCMS-C (ES-API): rt 2.22 min; m/z 749.3 [M+H]⁺. ¹H NMR (400 MHz, DMSO-d₆) δ 12.3 (s, 2H), 10.4 (s, 2H), 7.77-7.74 (m, 6H), 7.39-7.34 (m, 8H), 5.00-4.00 (m, 10H).

(c) 1,1′-(Ethane-1,2-diyl)bis(2-(2,4-difluoro-6-(5-oxo-4,5-dihydro-1,3,4-oxadiazol-2-yl)phenyl)-4-methoxy-1H-benzo[d]imidazole-5-carboxamide) (44)

To a solution of 1,1′-(ethane-1,2-diyl)bis(2-(2,4-difluoro-6-(hydrazinecarbonyl)phenyl)-4-methoxy-1H-benzo[d]imidazole-5-carboxamide) hydrochloride A47 (100 mg, assumed 0.10 mmol) in DMF (1 mL) was added CDI (64.9 mg, 0.40 mmol) and Et₃N (40.5 mg, 0.40 mmol) and the mixture was heated at 40° C. overnight. The mixture was concentrated under reduced pressure and the residue purified by prep. HPLC (Agilent YMC-C18, 5 μm, 250×20 mm column, eluting with a gradient of ACN in water with 0.1% Formic acid, at a flow rate of 15.0 mL/min) to give the title compound (2.5 mg, 3%) as a white solid. LCMS-B (ES-API): rt 4.08 min, m/z 801.0 [M+H]⁺. ¹H NMR (400 MHz, room temperature, DMSO-d₆) δ 12.7 (br s, 2H), 7.81-7.73 (m, 3H), 7.69-7.51 (m, 6H), 7.17 (d, J=8.4 Hz, 2H), 6.85 (s, 1H), 4.56 (d, J=9.6 Hz, 2H), 4.39 (s, 6H), 4.18 (d, J=9.6 Hz, 2H). ¹H NMR (400 MHz, 60° C., DMSO-d₆) δ 12.6 (s, 2H), 7.66-7.56 (m, 7H), 7.34 (s, 2H), 7.13 (d, J=8.8 Hz, 2H), 6.87 (s, 1H), 4.50 (s, 2H), 4.35 (s, 6H), 4.18 (s, 2H).

Example 22: 1,1-(Ethane-1,2-diyl)bis(2-(2-bromo-4-chloro-6-fluorophenyl)-4-methoxy-1H-benzo[d]imidazole-5-carboxamide) (45) and 1,1′-(ethane-1,2-diyl)bis(2-(4-chloro-2-fluoro-6-(2H-tetrazol-5-yl)phenyl)-4-methoxy-1H-benzo[d]imidazole-5-carboxamide) (46)

(a) 2-Bromo-4-chloro-6-fluorobenzaldehyde (A48)

To a solution of 1,2-dibromo-5-chloro-3-fluorobenzene (2.0 g, 6.94 mmol) in dry THF (10 mL) at −65° C. under N₂ was added i-PrMgCl (2 M solution in THF, 3.81 mL, 7.63 mmol) dropwise at such a rate that the temperature of the mixture did not exceed −50° C. The mixture was then cooled to −72° C. and stirred for 1 h. DMF (2.54 g, 34.6 mmol) was then added dropwise at such a rate that the temperature of the mixture did not exceed −55° C. and stirring was continued at −70° C. for 15 minutes. The mixture was quenched by dropwise addition of water (5 mL) and then extracted with EtOAc (50 mL×2). The combined organic extracts were washed with brine, dried over Na₂SO₄, filtered, and concentrated under reduced pressure and the residue was purified by silica gel chromatography (Petroleum Ether) to give the title compound (800 mg, 50%) as a white solid. ¹H NMR (400 MHz, CDCl₃) δ 10.20 (d, J=1.2 Hz, 1H), 7.45 (t, J=1.7 Hz, 1H), 7.13 (dd, J=10.0, 2.0 Hz, 1H).

(b) 1,1′-(Ethane-1,2-diyl)bis(2-(2-bromo-4-chloro-6-fluorophenyl)-4-methoxy-1H-benzo[d]imidazole-5-carboxamide) (45)

A mixture of 2-bromo-4-chloro-6-fluorobenzaldehyde (A48) (1.34 g, 5.67 mmol), 4,4′-(ethane-1,2-diylbis(azanediyl))bis(3-amino-2-methoxybenzamide) (I13) (1.0 g, 2.57 mmol) and NaHSO₃ (670 mg, 6.44 mmol) in DMF (16.8 mL) was heated at 80° C. overnight. The mixture was allowed to cool to room temperature, poured into water and the resulting precipitate collected by filtration. The collected solid was recrystallized from formic acid to give the title compound (700 mg, 33%) as a brown solid. LCMS-B (ES-API): rt 3.28 min, m/z 820.9, 822.9 [M+H]⁺.

(c) 1,1′-(Ethane-1,2-diyl)bis(2-(4-chloro-2-cyano-6-fluorophenyl)-4-methoxy-1H-benzo[d]imidazole-5-carboxamide) (A49)

A mixture of 1,1′-(ethane-1,2-diyl)bis(2-(2-bromo-4-chloro-6-fluorophenyl)-4-methoxy-1H-benzo[d]imidazole-5-carboxamide) (45) (700 mg, 0.85 mmol) and CuCN (228 mg, 2.55 mmol) in NMP (3.5 mL) was heated at 170° C. for 4 h. The mixture was allowed to cool to room temperature, poured into water and the resulting precipitate collected by filtration. The collected solid was purified by silica gel chromatography (DCM/MeOH, 100:1 to 10:1) to give the title compound (330 mg, 54%) as a brown solid. LCMS-C (ES-API): rt 3.49 min, m/z 715.06 [M+H]⁺.

(d) 1,1′-(Ethane-1,2-diyl)bis(2-(4-chloro-2-fluoro-6-(2H-tetrazol-5-yl)phenyl)-4-methoxy-1H-benzo[d]imidazole-5-carboxamide) (46)

A mixture of 1,1′-(ethane-1,2-diyl)bis(2-(4-chloro-2-cyano-6-fluorophenyl)-4-methoxy-1H-benzo[d]imidazole-5-carboxamide) (A49) (130 mg, 0.182 mmol), Et₃N.HCl (150 mg, 1.09 mmol) and NaN₃ (141.7 mg, 2.18 mmol) in DMF (2.4 mL) was heated at 80° C. under N₂ overnight. The mixture was returned to room temperature and poured into water. No precipitate formed so the mixture was concentrated under reduced pressure at 55° C. and the residue purified by silica gel chromatography (DCM/MeOH, 5:1 containing 2.5% v/v conc. aq. NH₄OH) followed by prep-HPLC (Agilent, YMC-C18, 150×20 mm, 5.0 μm column, eluting with a gradient of MeOH in water with 0.1% TFA at a flow rate of 15.0 mL/min) to give the title compound (14.7 mg, 11%) as a white solid. LCMS-B (ES-API): rt 2.68 min, m/z 801.2, 803.2 [M+H]⁺. ¹H NMR (400 MHz, DMSO-d₆) δ 7.96-7.63 (m, 6H), 7.58-7.33 (m, 4H), 7.24-7.09 (m, 2H), 4.84-4.55 (m, 2H), 4.43-4.25 (m, 2H), 4.14 (s, 6H).

Example 23: 2-(5-Carbamoyl-1-(2-(5-carbamoyl-2-(4-chloro-2-(2H-tetrazol-5-yl)phenyl)-4-methoxy-1H-benzo[d]imidazol-1-yl)ethyl)-4-methoxy-1H-benzo[d]imidazol-2-yl)-5-chloro-3-fluorobenzoic acid (47)

A mixture of 4,4′-(ethane-1,2-diylbis(azanediyl))bis(3-amino-2-methoxybenzamide) (I13) (100 mg, 0.26 mmol), 4-chloro-2-(2H-tetrazol-5-yl)benzaldehyde (A18) (54 mg, 0.26 mmol), 5-chloro-3-fluoro-2-formylbenzoic acid (A25) (52 mg, 0.26 mmol) and NaHSO₃ (80 mg, 0.78 mmol) in DMF (8.0 mL) was heated at 120° C. under a N₂ atmosphere for 16 h. The mixture was cooled to room temperature, poured into water (40 mL) and the resulting precipitate collected by filtration. Purification by prep-HPLC (Agilent 10 prep-C18, 10 μm, 250×21.3 mm column, eluting with a gradient of ACN in water with 0.1% formic acid, at a flow rate of 20 mL/min) gave the title compound (8.0 mg, 4%) as a white solid. LCMS-B (ES-API): rt 2.630 min, m/z 759.1, 761.2 [M+H]⁺. ¹H NMR (400 MHz, DMSO-d₆) δ 7.97 (d, J=1.6 Hz, 1H), 7.74-7.62 (m, 5H), 7.51-7.42 (m, 2H), 7.23-6.96 (m, 4H), 6.71-6.61 (m, 1H), 4.41-4.25 (m, 2H), 4.37 (s, 3H), 4.27 (s, 3H), 4.22-4.11 (m, 2H).

Example 24: 1,1-(Ethane-1,2-diyl)bis(2-(2-fluoro-6-(2H-tetrazol-5-yl)phenyl)-4-methoxy-1H-benzo[d]imidazole-5-carboxamide) (48)

(a) 2-(Bromomethyl)-3-fluorobenzonitrile (A50)

To a solution of 3-fluoro-2-methylbenzonitrile (4.4 g, 32.6 mmol) in CCl₄ (45 mL) was added NBS (29.0 g, 163 mmol) and BPO (0.8 g, 3.3 mmol) and the mixture was heated at reflux overnight. The mixture was returned to room temperature and the solids were removed by filtration and washed with EtOAc. The filtrate was concentrated under reduced pressure and the residue purified by silica gel chromatography (Pet. Ether/EtOAc=200:1 to 50:1) to give the title compound (5.2 g, 74%) as a white solid. ¹H NMR (400 MHz, DMSO-d₆) δ 7.82 (dd, J=7.6, 0.8 Hz, 1H), 7.70-7.60 (m, 2H), 4.74 (d, J=0.8 Hz, 2H).

(b) 3-Fluoro-2-formylbenzonitrile (A51)

To a solution of 2-(bromomethyl)-3-fluorobenzonitrile (A50) (1.0 g, 4.8 mmol) in acetonitrile (20 mL) was added NMO (2.3 g, 19.2 mmol) and the mixture was stirred at room temperature for 2 h. The mixture was poured into water, extracted with EtOAc and the organic extract dried over Na₂SO₄, filtered and concentrated under reduced pressure to give the title compound (540 mg, 75%) as a brown solid. ¹H NMR (400 MHz, DMSO-d₆) δ 10.3 (s, 1H), 7.94-7.85 (m, 2H), 7.85-7.77 (m, 1H).

(c) 1,1′-(Ethane-1,2-diyl)bis(2-(2-cyano-6-fluorophenyl)-4-methoxy-1H-benzo[d]imidazole-5-carboxamide) (A52)

To a solution of 3-fluoro-2-formylbenzonitrile (A51) (500 mg, 3.35 mmol) in DMF (5.0 mL) was added 4,4′-(ethane-1,2-diylbis(azanediyl))bis(3-amino-2-methoxybenzamide) (I13) (592 mg, 1.52 mmol) and NaHSO₃ (397 mg, 3.81 mmol) and the mixture was heated at 80° C. overnight. The mixture was concentrated under reduced pressure and the residue purified by silica gel chromatography (DCM/MeOH=150:1 to 30:1) to give the title compound (76 mg, 8%) as a brown solid. LCMS-A (ES-API): rt 2.8 min, m/z 647.2 [M+H]⁺. ¹H NMR (400 MHz, DMSO-d₆) δ 7.70 (br s, 2H), 7.66-7.64 (m, 4H), 7.55 (br s, 2H), 7.44-7.41 (m, 4H), 7.07 (d, J=8.7 Hz, 2H), 4.65 (s, 4H), 4.42 (s, 6H).

(d) 1,1′-(Ethane-1,2-diyl)bis(2-(2-fluoro-6-(2H-tetrazol-5-yl)phenyl)-4-methoxy-1H-benzo[d]imidazole-5-carboxamide) (48)

To a solution of 1,1′-(ethane-1,2-diyl)bis(2-(2-cyano-6-fluorophenyl)-4-methoxy-1H-benzo[d]imidazole-5-carboxamide) (A52) (76 mg, 118 mmol) in DMF (0.5 mL) was added Et₃N.HCl (97 mg, 0.705 mmol) and NaN₃ (92 mg, 1.42 mmol) and the mixture was heated at 80° C. overnight. The mixture was concentrated under reduced pressure and the residue purified by prep-HPLC (Agilent YMC-C18, 5 μm, 150×20 mm column, eluting with a gradient of ACN in water with 0.1% formic acid, at a flow rate of 15.0 mL/min) to give the title compound (4.6 mg, 5%) as a brown solid. LCMS-B (ES-API): rt 2.445 min; m/z 733.2 [M+H]⁺. ¹H NMR (400 MHz, DMSO-d₆) δ 8.03 (d, J=7.8 Hz, 1H), 7.83 (d, J=7.9 Hz, 2H), 7.70-7.58 (m, 4H), 7.56-7.37 (m, 4H), 6.96 (d, J=8.2 Hz, 2H), 6.49 (d, J=8.4 Hz, 1H), 4.40-4.25 (m, 2H), 4.19 (s, 6H), 4.09-3.99 (m, 2H).

Example 25: 1,1-(Ethane-1,2-diyl)bis(2-(2-fluoro-6-(5-oxo-4,5-dihydro-1,3,4-oxadiazol-2-yl)phenyl)-4-methoxy-1H-benzo[d]imidazole-5-carboxamide) (49)

(a) Methyl 2-bromo-3-fluorobenzoate (A53)

To a solution of 2-bromo-3-fluorobenzoic acid (5.00 g, 22.9 mmol) in MeOH (40 mL) was added conc. H₂SO₄ (3.0 mL) and the mixture was heated at reflux overnight. The mixture diluted with water, extracted with DCM and the organic layer washed with brine, dried over Na₂SO₄, filtered and concentrated under reduce pressure to give the title compound (4.8 g, 90%) as a white solid. ¹H NMR (400 MHz, DMSO-d₆) δ 7.71-7.49 (m, 3H), 3.88 (s, 3H).

(b) 2-Bromo-3-fluorobenzohydrazide (A54)

To a solution of methyl 2-bromo-3-fluorobenzoate (A53) (4.8 g, 20.6 mmol) in EtOH (30 mL) was added hydrazine hydrate (80% in water, 16.0 g, 329 mmol) and the mixture was heated at 55° C. for 3 h. The mixture was returned to room temperature, poured into ice water and the resulting precipitate collected by filtration and dried under vacuum to give the title compound (3.2 g, 67%) as a white solid. LCMS-B (ES-API): rt 1.54 min, m/z 233.0, 235.0 [M+H]⁺. ¹H NMR (400 MHz, DMSO-d₆) δ 9.64 (br s, 1H), 7.52-7.40 (m, 2H), 7.20 (ddd, J=6.9, 2.0, 0.8 Hz, 1H), 4.53 (br s, 2H).

(c) 5-(2-Bromo-3-fluorophenyl)-1,3,4-oxadiazol-2(3H)-one (A55)

To a solution of 2-bromo-3-fluorobenzohydrazide (A54) (3.2 g, 13.7 mmol) in DMF (26.0 mL) was added Et₃N (4.2 g, 41.2 mmol) and CDI (6.7 g, 41.2 mmol) and the mixture was heated at 40° C. for 3 h. The mixture was poured into water, extracted with DCM and the organic extract washed with brine, dried over Na₂SO₄, filtered and concentrated under reduced pressure. The residue was purified by silica gel chromatography (Pet. Ether/EtOAc=5:1) to give the title compound (1.5 g, 43%) as a white solid. ¹H NMR (400 MHz, DMSO-d₆) δ 12.9 (br s, 1H), 7.67-7.59 (m, 3H).

(d) 5-(3-Fluoro-2-vinylphenyl)-1,3,4-oxadiazol-2(3H)-one (A56)

To a solution of 5-(2-bromo-3-fluorophenyl)-1,3,4-oxadiazol-2(3H)-one (A55) (600.0 mg, 2.33 mmol) and tributyl(vinyl)stannane (2.2 g, 6.98 mmol) in toluene (9 mL) was added Pd(PPh₃)₄ (265.8 mg, 0.23 mmol) and the mixture was heated at reflux under N₂ for 16 h. The mixture was poured into water (20 mL), extracted with EtOAc (9 mL×3) and the combined organic extracts washed with water (9 mL×2), brine (9 mL), dried over Na₂SO₄, filtered and concentrated under reduced pressure. The residue was purified by silica gel chromatography (Pet. ether/EtOAc=50:1 to 10:1) to give the title compound (420 mg, 87%) as a white solid. LCMS-B (ES-API): rt 3.307 min, m/z 207.0 [M+H]⁺. ¹H NMR (400 MHz, DMSO-d₆) δ 12.7 (br s, 1H), 7.60-7.56 (m, 1H), 7.51-7.44 (m, 2H), 6.99 (dd, J=17.9, 11.8 Hz, 1H), 5.78 (ddd, J=17.9, 2.6, 1.6 Hz, 1H), 5.67 (dt, J=11.8, 1.8 Hz, 1H).

(e) 2-Fluoro-6-(5-oxo-4,5-dihydro-1,3,4-oxadiazol-2-yl)benzaldehyde (A57)

A solution of 5-(3-fluoro-2-vinylphenyl)-1,3,4-oxadiazol-2(3H)-one (A56) (200 mg, 0.97 mmol) in DCM/MeOH (6.0 mL/6.0 mL) was cooled to −70° C. and bubbled with O₃ gas until a pale blue color persisted. Dimethyl sulfide (746 mg, 12.0 mmol) was added, the mixture was returned to room temperature and then stirred for 1 h. The mixture was concentrated under reduced pressure to give the title compound (250 mg) as a yellow oil, which was used directly in the next step without further purification. LCMS-B (ES-API): rt 2.69 min, m/z 207.0 [M−H]⁻

(f) 1,1′-(Ethane-1,2-diyl)bis(2-(2-fluoro-6-(5-oxo-4,5-dihydro-1,3,4-oxadiazol-2-yl)phenyl)-4-methoxy-1H-benzo[d]imidazole-5-carboxamide) (49)

A solution of 2-fluoro-6-(5-oxo-4,5-dihydro-1,3,4-oxadiazol-2-yl)benzaldehyde (A57) (crude material from previous step, 250 mg), 4,4′-(ethane-1,2-diylbis(azanediyl))bis(3-amino-2-methoxybenzamide) (I13) (120 mg, 0.41 mmol) and Oxone® (304 mg, 0.49 mmol) in DMF (3.0 mL) was stirred at room temperature for 1 h. The mixture was poured into water (10.0 mL) and the resulting precipitate collected by filtration and purified by prep-HPLC (Agilent YMC-C18, 5 μm, 150×20 mm column, eluting with a gradient of ACN in water with 0.1% formic acid, at a flow rate of 15.0 mL/min) to give the title compound (9.0 mg, 2.4% over two steps) as a white solid. NMR analysis showed the presence of approximately 0.05 equivalents of formic acid. LCMS-B (ES-API): rt 2.63 min, m/z 765.2 [M+H]+; ¹H NMR (400 MHz, DMSO-d₆) δ 7.90-7.62 (m, 6H), 7.62-7.46 (m, 5H), 7.28-7.15 (m, 2H), 7.00-6.92 (m, 1H), 4.67-4.54 (m, 2H), 4.29 (s, 6H), 4.18-4.05 (m, 2H).

Example 26: 1,1-(Ethane-1,2-diyl)bis(2-(4-chloro-2-fluoro-6-(5-oxo-4,5-dihydro-1,3,4-oxadiazol-2-yl)phenyl)-4-methoxy-1H-benzo[d]imidazole-5-carboxamide) (50)

(a) Methyl 2-bromo-5-chloro-3-fluorobenzoate (A58)

To a solution of 2-bromo-5-chloro-3-fluorobenzoic acid (A24) (4.0 g, 15.8 mmol) in DMF (52 mL) was added CH₃I (4.5 g, 31.6 mmol) and K₂CO₃ (6.4 g, 47.4 mmol) and the mixture was stirred at room temperature for 30 min. The mixture was diluted with water, extracted with EtOAc and the organic layer washed with brine, dried over Na₂SO₄, filtered, and concentrated under reduced pressure. The residue was purified by silica gel chromatography (Pet. Ether/EtOAc=1:0 to 50:1) to give the title compound (3.97 g, 95%) as a white solid. ¹H NMR (400 MHz, DMSO-d₆) δ 7.87 (dd, J=8.6, 2.4 Hz, 1H), 7.70 (dd, J=2.3, 1.5 Hz, 1H), 3.88 (s, 3H).

(b) 2-Bromo-5-chloro-3-fluorobenzohydrazide (A59)

To a solution of methyl 2-bromo-5-chloro-3-fluorobenzoate (A58) (1.5 g, 5.6 mmol) in EtOH (10 mL) was added hydrazine hydrate (80% in water, 5.6 g, 89.5 mmol) and the mixture was heated at 55° C. for 2 h. The mixture was returned to room temperature, poured into ice water and the resulting precipitate collected by filtration and dried under vacuum to give the title compound as a white solid (1.4 g, 93%). ¹H NMR (400 MHz, DMSO-d₆) δ 9.71 (br s, 1H), 7.74 (dd, J=8.7, 2.4 Hz, 1H), 7.34 (dd, J=2.4, 1.3 Hz, 1H), 4.55 (br s, 2H).

(c) 5-(2-Bromo-5-chloro-3-fluorophenyl)-1,3,4-oxadiazol-2(3H)-one (A60)

To a solution of 2-bromo-5-chloro-3-fluorobenzohydrazide (A59) (1.4 g, 5.6 mmol) in DMF (15.0 mL) was added CDI (2.5 g, 15.7 mmol) and Et₃N (1.6 g, 15.7 mmol) and the mixture was heated at 40° C. overnight under N₂. The mixture was cooled to room temperature, poured into water and extracted with DCM. The organic extract was washed with brine, dried over Na₂SO₄, filtered and concentrated under reduced pressure. The residue was purified by silica gel chromatography (Pet. Ether/EtOAc=50:1 to 10:1) to give the title compound (630 mg, 42%) as a white solid. LCMS-B (ES-API): rt 3.543 min; m/z 292.9, 294.9 [M+H]⁺. ¹H NMR (400 MHz, DMSO-d₆) δ 7.90 (dd, J=8.6, 2.4 Hz, 1H), 7.70 (dd, J=2.0, 1.2 Hz, 1H).

(d) 5-(5-Chloro-3-fluoro-2-vinylphenyl)-1,3,4-oxadiazol-2(3H)-one (A61)

To a solution of 5-(2-bromo-5-chloro-3-fluorophenyl)-1,3,4-oxadiazol-2(3H)-one (A60) (630 mg, 2.17 mmol) in toluene (7.0 mL) was added tributyl(vinyl)stannane (2.06 g, 6.5 mmol) and Pd(PPh₃)₄ (375.3 mg, 0.325 mmol) and the mixture was heated at reflux under N₂ overnight. The mixture was concentrated under reduced pressure and the residue purified by silica gel chromatography (Pet. Ether to Pet. Ether/EtOAc=20:1) to give the title compound (380 mg, 73%) as a white solid. LCMS-B (ES-API): rt 3.656 min; m/z 241.0, 242.9 [M+H]⁺. ¹H NMR (400 MHz, DMSO-d₆) δ 12.9 (s, 1H), 7.76 (dd, J=10.9, 2.1 Hz, 1H), 7.76 (dd, J=2.0, 0.8 Hz, 1H), 6.98 (dd, J=18.0, 12.0 Hz, 1H), 5.82-5.68 (m, 2H).

(e) 4-Chloro-2-fluoro-6-(5-oxo-4,5-dihydro-1,3,4-oxadiazol-2-yl)benzaldehyde (A62)

A solution of 5-(5-chloro-3-fluoro-2-vinylphenyl)-1,3,4-oxadiazol-2(3H)-one (A61) (380 mg, 1.58 mmol) in DCM/MeOH (1:1, 5.0 mL) was cooled to −78° C. and bubbled with O₃ gas) until a pale blue color persisted. Dimethyl sulfide (490 mg, 7.9 mmol) was then added and the mixture was returned to room temperature and stirred for 1 h. The mixture was concentrated at 25° C. to give the title compound (200 mg) as a brown oil, which was used directly in the next step without further purification. LCMS-B (ES-API): rt 3.144 min; m/z 241.0, 243.0 [M−H]⁻.

(f) 1,1′-(Ethane-1,2-diyl)bis(2-(4-chloro-2-fluoro-6-(5-oxo-4,5-dihydro-1,3,4-oxadiazol-2-yl)phenyl)-4-methoxy-1H-benzo[d]imidazole-5-carboxamide) (50)

A solution of 4-chloro-2-fluoro-6-(5-oxo-4,5-dihydro-1,3,4-oxadiazol-2-yl) benzaldehyde (A62) (crude material from previous step, 200 mg), 4,4′-(ethane-1,2-diylbis(azane diyl))bis(3-amino-2-methoxybenzamide) (I13) (87.5 mg, 0.23 mmol) and Oxone® (170 mg, 0.276 mmol) in DMF (1.0 mL) was stirred at room temperature for 1 h. The mixture was then diluted with water, freeze-dried and the residue purified by silica gel chromatography (DCM to DCM/MeOH=10:1) followed by prep-HPLC (Agilent YMC-C18, 5 μm, 150×20 mm column, eluting with a gradient of ACN in water with 0.1% formic acid, at a flow rate of 15.0 mL/min) to give the title compound (7.2 mg, 1.1% over 2 steps) as a white solid. LCMS-B (ES-API): rt 2.799 min; m/z 833.1, 835.1 [M+11]+. ¹H NMR (400 MHz, DMSO-d₆) δ 7.73 (br s, 2H), 7.64 (s, 2H), 7.55-7.46 (m, 4H), 7.06 (d, J=8.6 Hz, 2H), 6.89 (br s, 2H), 4.50-4.39 (m, 2H), 4.33 (s, 6H), 4.15-4.04 (m, 2H).

Further Examples

The following example compounds were prepared by analogous methods.

Characterisation Structure Name data 19.1 and 19.2

2,2′-(Ethane-1,2-diylbis(5- carbamoyl-4-methoxy-1H- benzo[d]imidazole-1,2- diyl))bis(3-iodobenzoic acid) Mixture of atropisomers (meso and racemate) separated into two peaks (unassigned): Peak 1: 19.1 Peak 2: 19.2 19.1: LCMS-B (ES- API): rt 2.88 min, m/z 901.0 [M + H]⁺. 19.2: LCMS-B (ES- API): rt 3.07 min, m/z 901.0 [M + H]⁺. 20

6,6′-(Ethane-1,2-diylbis(5- carbamoyl-7-fluoro-4- methoxy-1H- benzo[d]imidazole-1,2- diyl))bis(3-chlorobenzoic acid) LCMS-B (ES- API): rt 2.77 min, m/z 753.3, 755.3 [M + H]⁺. 21

Dimethyl 6,6′-(ethane-1,2- diylbis(5-carbamoyl-4- methoxy-1H- benzo[d]imidazole-1,2- diyl))bis(2- bromobenzoate) LCMS-B (ES- API): rt 3.37 min, m/z 833.1, 835.1 [M + H]⁺. 22

2,2′-(Ethane-1,2-diylbis(5- carbamoyl-4-methoxy-1H- benzo[d]imidazole-1,2- diyl))bis(3-chlorobenzoic acid) Mixture of three atropisomers (meso and racemate) LCMS-B (ES- API): rt 2.75 & 2.92 min, m/z 717.1, 719.1 [M + H]⁺. 23

1,1′-(Ethane-1,2-diyl)bis(2- (2-chloro-6-(2H-tetrazol-5- yl)phenyl)-4-methoxy-1H- benzo[d]imidazole-5- carboxamide) Mixture of atropisomers separated (meso and racemate): Peak 2 (unassigned stereochemistry) LCMS-B (ES- API): rt 2.87 min, m/z 765.2, 767.2 [M + H]⁺. 24

1,1′-(Ethane-1,2-diyl)bis(2- (4-chloro-2-(2H-1,2,3- triazol-4-yl)phenyl)-4- methoxy-1H- benzo[d]imidazole-5- carboxamide) LCMS-B (ES- API): rt 2.77 min, m/z 763.4, 765.4 [M + H]⁺. 25

6,6′-(Ethane-1,2-diylbis(5- carbamoyl-4-methoxy-1H- benzo[d]imidazole-1,2- diyl))bis(3-methylbenzoic acid) LCMS-B (ES- API): rt 2.48 min, m/z 677.3 [M + H]⁺. 26

2,2′-(Ethane-1,2-diylbis(5- carbamoyl-4-methoxy-1H- benzo[d]imidazole-1,2- diyl))bis(4-methylbenzoic acid) LCMS-B (ES- API): rt 2.61 min, m/z 677.3 [M + H]⁺. 27

6,6′-(Ethane-1,2-diylbis(5- carbamoyl-4-methoxy-1H- benzo[d]imidazole-1,2- diyl))bis(3-fluorobenzoic acid) LCMS-B (ES- API): rt 2.48 min, m/z 685.2 [M + H]⁺. 28

2,2′-(Ethane-1,2-diylbis(5- carbamoyl-4-methoxy-1H- benzo[d]imidazole-1,2- diyl))bis(4-fluorobenzoic acid) LCMS-B (ES- API): rt 2.56 min, m/z 685.2 [M + H]⁺. 29.1 and 29.2

2,2′-(Ethane-1,2-diylbis(5- carbamoyl-1H- benzo[d]imidazole-1,2- diyl))bis(3-bromobenzoic acid) Mixture of atropisomers (meso and racemate) separated into two peaks (unassigned): Peak 1: 29.1 Peak 2: 29.2 29.1: LCMS-A (ES- API): rt 0.49 min, m/z 745.0, 747.0 [M + H]⁺. 29.2: LCMS-A (ES- API): rt 0.58 min, m/z 744.9, 746.9 [M + H]⁺. 30

3,3′-(Ethane-1,2-diylbis(5- carbamoyl-4-methoxy-1H- benzo[d]imidazole-1,2- diyl))diisonicotinic acid LCMS-B (ES- API): rt 3.09 min, m/z 651.2 [M + H]⁺. 31

2,2′-(Ethane-1,2-diylbis(5- carbamoyl-4-methoxy-1H- benzo[d]imidazole-1,2- diyl))bis(5-chloronicotinic acid) LCMS-B (ES- API): rt 2.85 min, m/z 719.3, 721.3 [M + H]⁺. 32

1,1′-(Ethane-1,2-diyl)bis(2- (4-chloro-2-(5-oxo-4,5- dihydro-1,2,4-oxadiazol-3- yl)phenyl)-4-methoxy-1H- benzo[d]imidazole-5- carboxamide) LCMS-B (ES- API): rt 2.77 min, m/z 797.4, 799.4 [M + H]⁺. 33

6,6′-(Ethane-1,2-diylbis(5- carbamoyl-4-methoxy-1H- benzo[d]imidazole-1,2- diyl))bis(3-ethylbenzoic acid) LCMS-B (ES- API): rt 2.75 min, m/z 705.4 [M + H]⁺. 34

(E)-6,6′-(But-2-ene-1,4- diylbis(5-carbamoyl-4- methoxy-1H- benzo[d]imidazole-1,2- diyl))bis(3-chlorobenzoic acid) LCMS-B (ES- API): rt 3.04 min, m/z 743.3, 745.3 [M + H]⁺. 35

6,6′-(Propane-1,3- diylbis(5-carbamoyl-4- methoxy-1H- benzo[d]imidazole-1,2- diyl))bis(3-chlorobenzoic acid) LCMS-B (ES- API): rt 2.92 min, m/z 731.3, 733.3 [M + H]⁺. 36

6,6′-(Ethane-1,2-diylbis(5- carbamoyl-4-methoxy-1H- benzo[d]imidazole-1,2- diyl))bis(2-bromobenzoic acid) LCMS-B (ES- API): rt 2.53 min, m/z 805.0, 807.0 [M + H]⁺. 37

1,1′-(Ethane-1,2-diyl)bis(2- (3,4-dichloro-2-(2H- tetrazol-5-yl)phenyl)-4- methoxy-1H- benzo[d]imidazole-5- carboxamide) LCMS-B (ES- API): rt 2.73 min, m/z 833.1, 835.1 [M + H]⁺. 38

6,6′-(Ethane-1,2-diylbis(5- carbamoyl-4-methoxy-1H- benzo[d]imidazole-1,2- diyl))bis(3- (trifluoromethyl)benzoic acid) LCMS-B (ES- API): rt 2.90 min, m/z 785.2 [M + H]⁺. 39

6,6′-(Ethane-1,2-diylbis(5- carbamoyl-1H- benzo[d]imidazole-1,2- diyl))bis(3-chlorobenzoic acid) LCMS-B (ES- API): rt 3.47 min, m/z 657.1, 659.1 [M + H]⁺. 40

1,1′-(Ethane-1,2-diyl)bis(2- (3-chloro-2-(2H-tetrazol-5- yl)phenyl)-4-methoxy-1H- benzo[d]imidazole-5- carboxamide) LCMS-B (ES- API): rt 2.58 min, m/z 765.2, 787.2 [M + H]⁺. 41

6,6′-(Ethane-1,2-diylbis(5- carbamoyl-4-methoxy-1H- benzo[d]imidazole-1,2- diyl))bis(3- methoxybenzoic acid) LCMS-B (ES- API): rt 2.36 min, m/z 709.2 [M + H]⁺. 51

2-(5-Carbamoyl-1-(2-(5- carbamoyl-2-(2-carboxy-4- cyanophenyl)-4-methoxy- 1H-benzo[d]imidazol-1 - yl)ethyl)-4-methoxy-1H- benzo[d]imidazol-2-yl)-5- cyanobenzoic acid LCMS-B (ES- API): rt 2.215 min, m/z 699.3 [M + H]⁺. 52

Methyl 2-(5-carbamoyl-1- (2-(5-carbamoyl-2-(4- cyano-2- (methoxycarbonyl)phenyl)- 4-methoxy-1H- benzo[d]imidazol-1- yl)ethyl)-4-methoxy-1H- benzo[d]imidazol-2-yl)-5- cyanobenzoate LCMS-B (ES- API): rt 2.817 min, m/z 727.4 [M + H]⁺.

Assays

Compounds disclosed herein are tested for activity as modulators of STING in the following assays. Each assay is designed to test different aspects of potential interaction of the compounds with STING. Therefore, compounds are considered active provided they demonstrate binding or modulation in any one (or more) of these assays.

Protein Production and Purification

Biophysical experiments were performed with three different recombinant human STING protein variants designated according to allelic nomenclature of Yi et al., (2013). Codon optimized DNA sequences (for expression in Escherichia coli) encoding amino acid residues 149 to 345 (Swiss Prot Q86WV6) of human STING (WT), human STING (HAQ) and human STING (R232H) were synthesised by GenScript USA Inc (Piscataway, N.J., USA). These were ligated into a modified pET43a E. coli expression vector designed to encode N-terminal His tag followed by tobacco etch virus protease (TEV) cleavage site and a STING gene sequence. The resulting protein sequences for the three allelic variants are listed below:

His-TEV-hSTING(WT) MGHHHHHHGTENLYFQGS E ¹⁴⁹KGNFNVAHGLAWSYYIGYLRLILPELQARIRTYNOHYNNL LRGAVSQRLYILLPLDCGVPDNLSMADPNIRFLDKLPQQTGDRAGIKDRVYSNSIYELLENG QRAGTCVLEYATPLQTLFAMSQYSQAGFSREDRLEQAKLFCRTLEDILADAPESQNNCRLI AYQEPADDSSFSLSQEVLRHLRQEEKEEVTVGS ³⁴⁵ His-TEV-hSTING(R232H) MGHHHHHHGTENLYFQGS E ¹⁴⁹SKGNFNVAHGLAWSYYIGYLRLILPELQARIRTYNQHYNNL LRGAVSQRLYILLPLDCGVPDNLSMADPNIRFLDKLPQQTGDHAGIKDRVYSNSIYELLENG QRAGTCVLEYATPLQTLFAMSQYSQAGFSREDRLEQAKLFCRTLEDILADAPESQNNCRLI AYQEPADDSSFSLSQEVLRHLRQEEKEEVTVGS ³⁴⁵ His-TEV-hSTING(HAQ) MGHHHHHHGTENLYFQGS E ¹⁴⁹KGNFNVAHGLAWSYYIGYLRLILPELQARIRTYNQHYNNL LRGAVSQRLYILLPLDCGVPDNLSMADPNIRFLDKLPQQTADRAGIKDRVYSNSIYELLENG QRAGTCVLEYATPLQTLFAMSQYSQAGFSREDRLEQAKLFCQTLEDILADAPESQNNCRLI AYQEPADDSSFSLSQEVLRHLRQEEKEEVTVGS ³⁴⁵

To produce recombinant human STING proteins, expression plasmid encoding above-described constructs were transformed into E. coli BL21 DE3 strain and grown with shaking at 37° C. in 2×1 L volumes of Terrific broth (TB) supplemented with 100 μg/ml Ampicillin until OD₆₀₀ of 0.8 was reached. Cultures were then cooled to 16° C. and protein expression induced by the addition of isopropyl β-D-1-thiogalactopyranoside to a final concentration of 0.5 mM and the cultures shaken overnight for further 16 hours. Following expression, cell cultures were centrifuged at 5000×g for 20 min and cell pellet stored frozen at −70° C.

Protein purification was initiated by thawing the cell pellet in Lysis buffer (25 mM Tris-HCl pH 8.0, 300 mM NaCl, 5 mM DTT, 2 mM MgCl₂, 10 mM imidazole, 0.5 mg/ml lysozyme, benzonase endonuclease [EMD Millipore], 1 mM PMSF, complete protease inhibitor tablets EDTA-free [Roche]) using a ratio of 7 ml of buffer per 1 g of cells. Cells were further lysed by 3 passes through an ice cooled Avestin C₅ cell crusher and then centrifuged at 48,000×g at 4° C. Supernatant (cell lysate) was filtered through a 5 μm filter and loaded onto 5 mL HiTrap IMAC Sepharose FF column (GE Healthcare) pre-equilibrated with IMAC wash buffer 1 (25 mM Tris-HCl pH 8.0, 300 mM NaCl, 5 mM DTT, 10 mM imidazole) using Profinia Affinity chromatography purification system (Bio-Rad). The IMAC column was then sequentially washed with IMAC Wash buffer 1 and IMAC Wash buffer 2 (25 mM Tris-HCl pH 8.0, 300 mM NaCl, 5 mM DTT, 20 mM Imidazole) and bound His-TEV-hSTING protein eluted with IMAC Elution buffer (25 mM Tris-HCl pH 8.0, 300 mM NaCl, 5 mM DTT, 250 mM Imidazole). IMAC-eluted protein was further purified by passing through a HiLoad 26/60 Superdex 75 column pre-equilibrated in storage buffer (25 mM Tris-HCl, pH 8.0, 150 mM NaCl, 5 mM DTT 0.02% [w/v] sodium azide). Finally, hSTING protein was concentrated to 2 mg/ml using Amicon Ultra centrifugal filter unit (Utra-15 MWCO 10 kDa), flash-frozen in liquid nitrogen and stored in −70° C. freezer.

Differential Scanning Fluorimetry (DSF)

Differential scanning fluorimetry (DSF) is a rapid screening method for identifying low-molecular-weight ligands that bind and, in doing so, stabilize (or sometimes destabilize) purified proteins (Niesen 2007). DSF monitors thermal unfolding of protein in the presence of a fluorescent dye and is typically performed using a real-time PCR instrument. The temperature at which a protein unfolds is measured by an increase in the fluorescence of a dye with affinity for hydrophobic parts of the protein that are gradually being exposed during unfolding. The fluorescence of the dye is quenched in aqueous environments, but when the dye associates with hydrophobic sites on unfolded protein, its fluorescence increases. The fluorescence intensity is plotted as a function of the temperature, generating a sigmoidal curve that can be described by a two-state transition. The inflection point of the transition curve (Tm) is calculated using simple equations such as that of Boltzmann.

Thermal stability of STING protein (with and without bound ligand) was measured using previously described methodology (Seabrook & Newman, 2013), with samples tested in triplicate using 96-well PCR microplate (AB Gene, ABGAB-0600/W). In a final volume of 20 μL, 2 μM protein in 1×HBS buffer (50 mM HEPES, pH 7.4, 150 mM NaCl) was mixed with SYPRO Orange dye (Sigma-Aldrich S5692, final reaction mix dilution 1:1200) and a compound (final concentration at 100 μM). Sealed plates were placed into Bio-Rad CFX96/C1000 thermocycler and FRET scanning mode (λ^(excitation) of 490 nm and reads at a λ^(emission) of 570 nm). Melting curves were recorded from 20° C. to 100° C. in 0.5° C. increments every 30 seconds with a read at each increment. Data were analysed using “Meltdown analysis” protocol described by Rosa (2015). The melting temperature (T_(m)) obtained for STING protein alone (1%[v/v] DMSO) was subtracted from T_(m) obtained for protein incubated with ligand to generate ΔT_(m) values listed in the table below. DSF data was generated for each compound against 3 different STING protein variants—human STING (WT), human STING (HAQ) and human STING (R232H).

DSF huSTING DSF huSTING (HAQ) (WT) Example ΔTm (° C.) ΔTm (° C.)  1 9.44 5.07  2 −3.48 −6.94  3 2.66 −0.02  4 7.23 3.28  5 6.04 1.33  6 5.44 0.94  7 5.88 −2.49  8 23.22 6.38  9 11.31 5.06 10 18.43 9.63 11 22.12 10.62 12 28.33 4.56 14 9.15 4.74 15 4.78 −0.91 16 17.24 7.95 17 11.95 3.05 18 19.08 2.40 19.1 31.74 19.29 19.2 21.66 6.58 20 3.69 −3.00 21 5.83 2.33 22 28.16 16.96 23 29.21 14.96 24 −0.08 −3.16 25 −15.64 −18.45 26 6.33 1.13 27 6.17 2.62 28 −2.19 −6.60 29.1 7.46 −0.07 29.2 −5.65 −6.70 30 2.14 0.29 31 3.98 0.18 32 3.99 0.51 33 4.06 1.17 34 0.49 −2.39 35 1.28 −2.51 36 5.24 2.04 37 2.80 −0.24 38 3.68 0.38 39 0.86 −4.35 40 −1.74 −7.45 41 4.52 1.09 42 6.3 2.04 43 22.22 9.87 44 13.05 0.88 45 1.24 −0.34 46 28.61 14.38 48 25.49 11.8 49 21.32 10.22 50 26.42 12.47 51 1.6 −0.85

Surface Plasmon Resonance (SPR)

Binding interactions of ligands with STING proteins were quantified using Surface Plasmon Resonance (SPR) with a minimally biotinylated STING protein immobilized on a streptavidin chip surface. In this manner highly active STING protein surfaces were obtained that were not compromised by a low pH required for an amine coupling method. Minimal biotinylation of purified huSTING proteins was performed using a previously described methodology (Chhabra 2012). Briefly, approximately 20 nmol of recombinant STING protein in 1×TBS buffer (25 mM Tris-HCl, pH 7.5, 150 mM NaCl, 5 mM DTT) was mixed with of EZ-Link™ Sulfo-NHS-LC-LC-Biotin (Thermofisher Scientific, cat #21338) at a molar ratio of 1 to 0.6 and incubated on ice for 2 hours. To remove any unreacted biotin reagent, protein/biotin mixture was passed through a Superdex 75 (10/300 GL) column equilibrated with 10 mM HEPES, pH7.4, 150 mM NaCl, 5 mM DTT, 5%[v/v] glycerol. A protein peak containing biotinylated huSTING protein was collected and stored in aliquots at −80° C.

Streptavidin was simultaneously immobilized in all four channels of a CM5 sensor chip docked in a Biacore instrument (either Biacore S200 or Biacore T200, GE Healthcare) as described previously (Zender 2013). Minimally biotinylated STING protein was captured onto a streptavidin coated chip surface at 8° C. in SPR binding buffer (50 mM HEPES, pH 7.4, 150 mM NaCl, 2%[v/v] DMSO) by gradually injecting in a single channel at a constant flow-rate of 2 μL/min until desired capture level was achieved, typically 3000 to 7000 RU (1 RU=1 pg/mm²).

All binding experiments were performed at 8° C. in SPR binding buffer. To determine binding affinity, compound interaction with immobilized STING protein was analysed using dose-response experiments. Fresh 10 mM DMSO solutions of compound were diluted directly into SPR binding buffer typically to a concentration of 50 μM and then further diluted 2-fold or 3-fold aiming for either a 5- or 7-point concentration series range. Each ligand concentration series was injected at a constant flow rate of 60 μL/min with a 90 second association and a 180 second dissociation time. These were modified for compounds with longer residence times, so that curves could reach steady-state, or so that compound would be fully dissociated before the subsequent injection. Where appropriate, tighter-binding compounds (roughly K_(D)<1 uM) were tested using a single-cycle kinetics format (Karlsson 2006), with long association and dissociation times (typically 450 s and 1800 s, respectively).

Scrubber 2 (www.biologic.com.au) was utilized for data processing, where signals were referenced against the blank surface (streptavidin+D-biotin) and further corrected for DMSO refractive index change and then “double-referenced” using a buffer-blank injection (Papalia 2006). Responses were fitted to either a 1:1 steady-state affinity model or a 1:1 kinetic model (that included a mass-transport component), available within Scrubber.

SPR (HAQ) K_(D) Example (μM) 1 16.6 2 67 3 >10 4 28.6 5 19.3 6 11.9 9 2.66 10 0.183 11 0.078 15 6.34 16 0.166 17 0.416 18 0.144 19.1 0.002 20 12.8 21 1.09 22 0.002 23 0.002 24 22 25 19.3 26 32.5 29.1 0.936 30 90 31 55 32 90 33 78 34 26 35 35 37 0.200 38 >100 39 300 40 64 41 83 42 14.8 43 0.096 44 0.423 45 28.9 46 0.025 48 0.030 49 0.031 50 0.014 SPR (WT) K_(D) Example (μM) 1 200 2 80 3 700 4 >10 5 30 6 40 9 40 10 5.86 11 1.08 15 1000 16 1.52 17 9.5 18 2.01 19.1 0.034 20 96 21 5.380 22 0.077 23 0.043 24 102 25 250 26 >40 29.1 33 30 126 31 88 32 >80 33 107 34 74 35 >100 36 58 38 >100 40 180 41 200 42 139.1 43 1.79 44 11.1 45 29.6 46 0.413 48 0.512 49 0.755 50 0.359

THP-1 Reporter Cell Line Assays

THP-1 STING Lucia ISG cells (InvivoGen #thpI-isg) were cultured in RPMI-1640 containing 2 mM L-glutamine, 25 mM HEPES, 100 μg/mL Normocin (InvivoGen) and 10% heat-inactivated FBS. Cells were seeded at a density of 7×10⁵ cells/mL, maintained at 37° C./5% CO₂, and passaged every 3-4 days. Selection pressure was maintained by the addition of 100 μg/mL Zeocin every second passage.

Assay conditions: cells were harvested and resuspended at a concentration of 5×10⁵ cells/mL in fresh growth media. Of this cell preparation 20 μL per well was dispensed into 384 well cell culture plates (Greiner, #781098X) and incubated at 37° C./5% CO₂ for 2 hours. Concentration Response Curves (commonly 11 doses) were prepared by 2.5 fold serial dilution starting from 10 mM compound stock solution in DMSO. Compound dilutions were transferred into the cell culture plates using a pintool (0.1 μL transfer), control wells were matched for DMSO. Positive control wells received ML RR-S2 CDA (Med Chem Express HY12885B) in 5 μL of media to a final concentration of 40 μM. All other wells received additional 5 μL of media only to equalise volume. The plates were incubated for 24 h at 37° C./5% CO₂.

For the detection step, QUANTI-Luc (InvivoGen #rep-qlc1) was prepared according to manufacturer's instruction and 10 μL were added per well to the culture plates. Plates were shaken for ten seconds on an orbital shaker and left in the dark at room temperature for two minutes, before luminescence was read on a Perkin Elmer Envision plate reader.

Data analysis: Luminescence raw data was normalised % activity relative to the signal of the positive control wells and negative control wells which were not treated with compound. The following formula was used to calculate normalised % activity from raw signals:

% activity=(sample−negative)/(positive−negative)

The THP-1 STING Lucia ISG cell EC₅₀ provides an assessment of activity at the human HAQ isoform of STING. For assessing compound activity against the WT human STING isoform, an equivalent method was conducted where the cells used were THP-1-Dual KI-hSTING-R232 (InvivoGen #thpd-r232).

EC₅₀ THP-1 EC₅₀ THP-1 STING Lucia Dual KI-hSTING- ISG cells R232 cells Example (μM) (μM) 1 1.07 0.895 2 >40 >40 3 >40 >40 4 15.9 32.5 5 24.8 6.4 6 0.902 0.936 7 1.55 1.43 8 0.540 0.426 9 0.777 0.715 10 1.59 1.39 11 0.830 0.683 12 2.65 2.55 13 >40 >40 14 1.31 1.99 15 2.56 2.46 16 0.738 0.840 17 0.290 0.360 18 0.326 0.566 19.1 0.297 0.232 19.2 14.9 11.7 20 1.30 2.78 21 1.49 1.63 22 1.53 1.45 23 2.50 2.47 24 2.67 4.19 25 2.96 3.20 26 3.38 3.22 27 11.25 10.52 28 12.6 >40 29.1 18.4 9.85 29.2 >40 >40 30 39.1 28.9 31 >40 >40 32 >40 29.7 33 >40 >40 34 >40 >40 35 >40 >40 36 >40 >40 37 >40 >40 38 >40 >40 39 >40 >40 40 >40 >40 41 >40 >40 42 3.211 4.385 43 0.5743 0.4401 44 0.5576 0.3797 45 6.788 3.248 46 0.2175 0.1946 47 0.497 0.5493 48 0.4148 0.2694 49 0.136 0.1905 50 0.1736 0.03117

STATEMENTS OF INVENTION

1. A compound of formula I:

wherein: Y is either (CH₂)_(n), where n is from 2 to 4, or —CH₂—CH═CH—CH₂—; R¹ and R¹¹ are independently selected from the group consisting of: —C(═O)OH, —C(═O)OR^(P1), tetrazolyl, Br, F, oxo-oxadiazolyl and

(2H-triazol-4-yl), where R^(P1) is selected from methyl and ethyl; A¹ is CR^(A) or N; A² is CR^(B) or N; A³ is CR^(C) or N; A⁴ is CR^(D) or N; where no more than two of A¹, A², A³, and A⁴ may be N; one, two or three of R^(A), R^(B), R^(C) and R^(D), (if present) are selected from H, F, Cl, Br, I, Me, Et, CF₃, cyclopropyl, cyano, OMe, OEt, CH₂OH, CH₂OMe and OH; the remainder of R^(A), R^(B), R^(C) and R^(D), (if present) are H; A¹¹ is CR^(AA) or N; A¹² is CR^(BB) or N; A¹³ is CR^(CC) or N; A¹⁴ is OR^(DD) or N; where no more than two of A¹¹, A¹², A¹³ and A¹⁴ may be N; one, two or three of R^(AA), R^(BB), R^(CC) and R^(DD) (if present) are selected from H, F, Cl, Br, I, Me, Et, CF₃, cyclopropyl, cyano, OMe, OEt, CH₂OH, CH₂OMe and OH; the remainder of R^(AA), R^(BB), R^(CC) and R^(DD), (if present) are H; R^(C1), R^(C3) and R^(C4) are independently selected from H, Cl, F, Br, Me, OMe, OEt, cyano, CF₃, CH₂OH, CH₂OMe, C₂₋₄ alkenyl and C₅heterocyclyl; R^(C11), R^(C13) and R^(C14) are independently selected from H, Cl, F, Br, Me, OMe, OEt, cyano, CF₃, CH₂OH, CH₂OMe, C₂₋₄alkenyl and C₅heterocyclyl.

2. A compound according to statement 1, wherein A¹¹=A1, A¹²=A², A¹³=A³, A¹⁴=A⁴, R^(C11)=R^(C1), R^(C13)=R^(C3), R^(C14)=R^(C4), R¹¹=R¹.

3. A compound according to statements 1 or 2, wherein A¹ is CR^(A). 4. A compound according to statements 1 or 2, wherein A¹ is N.

5. A compound according to any one of statements 1 to 4, wherein A² is CR^(B).

6. A compound according to any one of statements 1 to 4, wherein A² is N.

7. A compound according to any one of statements 1 to 6, wherein A³ is CR^(C).

8. A compound according to any one of statements 1 to 6, wherein A³ is N.

9. A compound according to any one of statements 1 to 6, wherein A⁴ is CR^(D).

10. A compound according to any one of statements 1 to 6, wherein A⁴ is N.

11. A compound according to statements 1 or 2, wherein A¹ is CR^(A), A² is CR^(B), A³ is CR^(C), and A⁴ is CR^(D).

12. A compound according to statements 1 or 2, wherein one of A¹, A², A³ and A⁴ are N.

13. A compound according to statements 1 or 2, wherein two of A¹, A², A³ and A⁴ are N.

14. A compound according to any one of statements 1 to 13, wherein A¹¹ is CR^(AA).

15. A compound according to any one of statements 1 to 13, wherein A¹¹ is N.

16. A compound according to any one of statements 1 to 15, wherein A¹² is CR^(BB).

17. A compound according to any one of statements 1 to 15, wherein A¹² is N.

18. A compound according to any one of statements 1 to 17, wherein A¹³ is CR^(CC).

19. A compound according to any one of statements 1 to 17, wherein A¹³ is N.

20. A compound according to any one of statements 1 to 19, wherein A¹⁴ is CR^(D)D.

21. A compound according to any one of statements 1 to 19, wherein A¹⁴ is N.

22. A compound according to any one of statements 1 to 13, wherein A¹¹ is CR^(AA), A² is CR^(BB), A¹³ is CR^(CC), and A¹⁴ is CR^(D)D.

23. A compound according to any one of statements 1 to 13, wherein one of A¹¹, A¹², A¹³ and A¹⁴ are N.

24. A compound according to any one of statements 1 to 13, wherein two of A¹¹, A¹², A¹³ and A¹⁴ are N.

25. A compound according to any one of statements 1 to 13, wherein the compound is of formula IIIb:

26. A compound according to any one of statements 1 to 13, wherein the compound is of formula IIIc:

27. A compound according to any one of statements 1 to 13, wherein the compound is of formula IIId:

28. A compound according to any one of statements 1 to 13, wherein the compound is of formula IIIe:

29. A compound according to any one of statements 1 to 28, wherein one, two or three of R^(A), R^(B), R^(C) and R^(D), (if present) are selected from H, F, Cl, Br, Me, CF₃, cyclopropyl, cyano and OMe, the remainder (if present) are H.

30. A compound according to any one of statements 1 to 28, wherein one, two or three of R^(A), R^(B), R^(C) and R^(D), (if present) are selected from H, F, Cl, Br, Me, CF₃, cyclopropyl and OMe, the remainder (if present) are H.

31. A compound according to any one of statements 1 to 28, wherein one, two or three of R^(A), R^(B), R^(C) and R^(D), (if present) are selected from H, F, Cl, Br, Me, CF₃ and OMe, the remainder (if present) are H.

32. A compound according to any one of statements 1 to 31, wherein R^(A) (if present) is selected from H and F.

33. A compound according to any one of statements 1 to 32, wherein R^(B) (if present) is selected from H, F and Cl.

34. A compound according to statement 33, wherein R^(B) (if present) is selected from H and Cl.

35. A compound according to any one of statements 1 to 34, wherein R^(C) (if present) is selected from H, F and Cl.

36. A compound according to any one of statements 1 to 35, wherein R^(D) (if present) is selected from H, F and Cl.

37. A compound according to statement 11, wherein R^(A), R^(B), R^(C) and R^(D) are selected from one of the following combinations:

Combination R^(A) R^(B) R^(C) R^(D)  1 F H H H  2 H Br H H  3 H Cl H H  4 F Cl H H  5 H H H Br  6 H H Cl H  7 H F F H  8 H H H H  9 CF CH CF CH 10 CF CH CCl CH 11 CBr CH CH CH 12 CCl CH CH CH 13 N CH CCl CH 14 CH CCl CCl CH 15 CH CF CH CH 16 CH CMe CH CH 17 CH N CH CH 18 CH CH CBr CH 19 CH CH CCF₃ CH 20 CH CH CEt CH 21 CH CH CF CH 22 CH CH CMe CH 23 CH CH C—OMe CH 24 CH CH CCl CCl 25 CH CH CH CF 26 CH CH CH C—OMe 27 CH CH CH N 28 Cl CH CH CH 29 CH CH CCN CH 30 CBr CH CCl CH

38. A compound according to statement 37, wherein R^(A), R^(B), R^(C) and R^(D) are selected from combinations 1, 3, 6 and 9.

39. A compound according to any one of statements 1 to 38, wherein one, two or three of R^(AA), R^(BB), R^(CC) and R^(DD), (if present) are selected from H, F, Cl, Br, Me, CF₃, cyclopropyl, cyano and OMe, the remainder (if present) are H.

40. A compound according to any one of statements 1 to 38, wherein one, two or three of R^(AA), R^(BB), R^(CC) and R^(DD), (if present) are selected from H, F, Cl, Br, Me, CF₃, cyclopropyl and OMe, the remainder (if present) are H.

41. A compound according to any one of statements 1 to 38, wherein one, two or three of R^(AA), R^(BB), R^(CC) and R^(DD), (if present) are selected from H, F, Cl, Br, Me, CF₃ and OMe, the remainder (if present) are H.

42. A compound according to any one of statements 1 to 41, wherein R^(AA) (if present) is selected from H and F.

43. A compound according to any one of statements 1 to 42, wherein R^(BB) (if present) is selected from H, F and Cl.

44. A compound according to statement 43, wherein R^(BB) (if present) is selected from H and Cl.

45. A compound according to any one of statements 1 to 44, wherein R^(CC) (if present) is selected from H, F and Cl.

46. A compound according to any one of statements 1 to 45, wherein R^(DD) (if present) is selected from H, F and Cl.

47. A compound according to statement 22, wherein R^(AA), R^(BB), R^(CC) and R^(DD) are selected from one of the following combinations:

Combination R^(AA) R^(BB) R^(CC) R^(DD)  1 F H H H  2 H Br H H  3 H Cl H H  4 F Cl H H  5 H H H Br  6 H H Cl H  7 H F F H  8 H H H H  9 CF CH CF CH 10 CF CH CCl CH 11 CBr CH CH CH 12 CCl CH CH CH 13 N CH CCl CH 14 CH CCl CCl CH 15 CH CF CH CH 16 CH CMe CH CH 17 CH N CH CH 18 CH CH CBr CH 19 CH CH CCF₃ CH 20 CH CH CEt CH 21 CH CH CF CH 22 CH CH CMe CH 23 CH CH C—OMe CH 24 CH CH CCl CCl 25 CH CH CH CF 26 CH CH CH C—OMe 27 CH CH CH N 28 Cl CH CH CH 29 CH CH CCN CH 30 CBr CH CCl CH

48. A compound according to statement 47, wherein R^(AA), R^(BB), R^(CC) and R^(DD) are selected R^(BB), from combinations 1, 3, 6 and 9.

49. A compound according to any one of statements 1 to 48, wherein R^(C1), R^(C3) and R^(C4) are independently selected from H, Cl, F, Br, Me, OMe, cyano, CF₃ and CH₂OH.

50. A compound according to any one of statements 1 to 49, wherein R^(C1), R^(C3) and R^(C4) are independently selected from H, Cl, F, CF₃, OMe and CH₂OH

51. A compound according to any one of statements 1 to 50, wherein R^(C1), R^(C3) and R^(C4) are independently selected from H, F and OMe.

52. A compound according to any one of statements 1 to 51, wherein two of R^(C1), R^(C3) and R^(C4) are H, and the other is selected from the defined groups, except H.

53. A compound according to any one of statements 1 to 52, wherein one of R^(C1), R^(C3) and R^(C4) is H, and the other two are independently selected from the defined groups, except H.

54. A compound according to any one of statements 1 to 52, wherein R^(C1), R^(C3) and R^(C4) are H.

55. A compound according to any one of statements 1 to 48, wherein:

-   -   R^(C1) is H;     -   R^(C3) is H; and     -   R^(C4) is F.

56. A compound according to any one of statements 1 to 48, wherein:

-   -   R^(C1) is OMe;     -   R^(C3) is H; and     -   R^(C4) is H or F.

57. A compound according to any one of statements 1 to 56, wherein R^(C11)=R^(C1), R^(C13)=R^(C3) and R^(C14)=R^(C4).

58. A compound according to any one of statements 1 to 56, wherein R^(C11), R^(C13) and R^(C14) are independently selected from H, Cl, F, Br, Me, OMe, cyano, CF₃ and CH₂OH.

59. A compound according to any one of statements 1 to 56, wherein R^(C11), R^(C13) and R^(C14) are independently selected from H, Cl, F, CF₃, OMe and CH₂OH.

60. A compound according to any one of statements 1 to 56, wherein R^(C11), R^(C13) and R^(C14) are independently selected from H, F and OMe.

61. A compound according to any one of statements 1 to 60, wherein two of R^(C11), R^(C13) and R^(C14) are H, and the other is selected from the defined groups, except H.

62. A compound according to any one of statements 1 to 60, wherein one of R^(C11), R^(C13) and R^(C14) is H, and the other two are independently selected from the defined groups, except H.

63. A compound according to any one of statements 1 to 60, wherein R^(C11), R^(C13) and R^(C14) are H.

64. A compound according to any one of statements 1 to 56, wherein:

-   -   R^(C11) is H;     -   R^(C13) is H; and     -   R^(C14) is F.

65. A compound according to any one of statements 1 to 56, wherein:

-   -   R^(C11) is OMe;     -   R^(C13) is H; and     -   R^(C14) is H.

66. A compound according to any one of statements 1 to 65, wherein Y is (CH₂)_(n), where n is from 2 to 3.

67. A compound according to any one of statements 1 to 65, wherein Y is (CH₂)₂.

68. A compound according to any one of statements 1 to 65, wherein Y is (CH₂)₃.

69. A compound according to any one of statements 1 to 65, wherein Y is (CH₂)₄.

70. A compound according to any one of statements 1 to 65, wherein Y is —CH₂—CH═CH—CH₂ ⁻.

71. A compound according to any one of statements 1 to 70, wherein R¹ and R¹¹ are selected from —C(═O)OH, —C(═O)OR^(P1), tetrazolyl, oxo-oxadiazolyl and

(2H-triazol-4-yl), where R^(P1) is selected from methyl and ethyl.

72. A compound according to any one of statements 1 to 71, wherein R¹ and R¹¹ are selected from C(═O)OH, tetrazolyl and oxo-oxadiazolyl.

73. A compound according to statement 71 or 72, wherein R¹ and R¹¹ are C(═O)OH.

74. A compound according to statement 71 or 72, wherein R¹ and R¹¹ are tetrazolyl

75. A compound according to statement 71 or 72, wherein R¹ and R¹¹ are oxo-oxadiazolyl.

76. A compound according to any one of statements 1 to 76, wherein the compound is selected from any one of compounds 1-52.

77. A compound according to any one of statements 1-75, wherein the compound is selected from any one of compounds 1-51.

78. A compound according to any one of statements 1-77, wherein the compound is in the form of a pharmaceutically acceptable salt, solvate, prodrug, isomer, tautomer, polymorph or N-oxide, or a combination thereof in any ratio.

79. A compound as defined in any one of statements 1 to 78, for use in a method of therapy.

80. A pharmaceutical composition comprising a compound as defined in any one of statements 1 to 78, and a pharmaceutically acceptable excipient.

81. A method of treatment or prevention of a disease ameliorated by the modulation of STING, comprising administering to a patient in need of treatment, a compound as defined in any one of statements 1 to 78, or a pharmaceutical composition according to statement 80.

82. The use of a compound as defined in any one of statements 1 to 78, in the manufacture of a medicament for treating or preventing disease ameliorated by the modulation of STING.

83. A compound as defined in any one of statements 1 to 78, or pharmaceutical composition according to statement 78 for use in the treatment or preventing of disease ameliorated by the modulation of STING.

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1. A compound of formula I:

wherein: Y is either (CH₂)_(n), where n is from 2 to 4, or —CH₂—CH═CH—CH₂—; R¹ and R¹¹ are independently selected from —C(═O)OH, —C(═O)OR^(P1), Br, F, tetrazolyl, oxo-oxadiazolyl and

 (2H-triazol-4-yl), where R^(P1) is selected from methyl and ethyl; A¹ is CR^(A) or N; A² is CR^(B) or N; A³ is CR^(C) or N; A⁴ is CR^(D) or N; where no more than two of A¹, A², A³, and A⁴ may be N; one, two or three of R^(A), R^(B), R^(C) and R^(D), (if present) are selected from H, F, Cl, Br, I, Me, Et, CF₃, cyclopropyl, cyano, OMe, OEt, CH₂OH, CH₂OMe and OH; the remainder of R^(A), R^(B), R^(C) and R^(D), (if present) are H; A¹¹ is CR^(AA) or N; A¹² is CR^(BB) or N; A¹³ is CR^(CC) or N; A¹⁴ is CR^(DD) or N; where no more than two of A¹¹, A¹², A¹³ and A¹⁴ may be N; one, two or three of R^(AA), R^(BB), R^(CC) and R^(DD) (if present) are selected from H, F, Cl, Br, I, Me, Et, CF₃, cyclopropyl, cyano, OMe, OEt, CH₂OH, CH₂OMe and OH; the remainder of R^(AA), R^(BB), R^(CC) and R^(DD), (if present) are H; R^(C1), R^(C3) and R^(C4) are independently selected from H, Cl, F, Br, Me, OMe, OEt, cyano, CF₃, CH₂OH, CH₂OMe, C₂₋₄ alkenyl and C₅heterocyclyl; R^(C11), R^(C13) and R^(C14) are independently selected from H, Cl, F, Br, Me, OMe, OEt, cyano, CF₃, CH₂OH, CH₂OMe, C₂₋₄alkenyl and C₅heterocyclyl; or a pharmaceutically acceptable salt, solvate, prodrug, isomer, tautomer, polymorph and/or N-oxide thereof.
 2. A compound according to claim 1, or a pharmaceutically acceptable salt, solvate, prodrug, isomer, tautomer, polymorph and/or N-oxide thereof, wherein A¹¹=A¹, A¹²=A², A¹³=A³, A¹⁴=A⁴, R^(C13)=R^(C3), R^(C14)=R^(C4), R¹¹=R¹.
 3. A compound according to either claim 1 or claim 2, or a pharmaceutically acceptable salt, solvate, prodrug, isomer, tautomer, polymorph and/or N-oxide thereof, wherein A¹ is CR^(A), A² is CR^(B), A³ is CR^(C), and A⁴ is CR^(D).
 4. A compound according to any one of claims 1 to 3, or a pharmaceutically acceptable salt, solvate, prodrug, isomer, tautomer, polymorph and/or N-oxide thereof, wherein A¹¹ is CR^(AA), A¹² is CR^(BB), A¹³ is CR^(CC), and A¹⁴ is CR^(DD).
 5. A compound according to either claim 1 or claim 2, or a pharmaceutically acceptable salt, solvate, prodrug, isomer, tautomer, polymorph and/or N-oxide thereof, wherein the compound is selected from formulae IIIb, IIIc, IIId and IIIe:


6. A compound according to any one of claims 1 to 5, or a pharmaceutically acceptable salt, solvate, prodrug, isomer, tautomer, polymorph and/or N-oxide thereof, wherein one, two or three of R^(A), R^(B), R^(C) and R^(D), (if present) are selected from H, F, Cl, Br, Me, CF₃, cyclopropyl, cyano and OMe, the remainder (if present) are H.
 7. A compound according to any one of claims 1 to 4, or a pharmaceutically acceptable salt, solvate, prodrug, isomer, tautomer, polymorph and/or N-oxide thereof, wherein A¹, A², A³ and A⁴ are selected from combinations 1-30: Combination A¹ A² A³ A⁴  1 CF CH CH CH  2 CH CBr CH CH  3 CH CCl CH CH  4 CF CCl CH CH  5 CH CH CH CBr  6 CH CH CCl CH  7 CH CF CF CH  8 CH CH CH CH  9 CF CH CF CH 10 CF CH CCl CH 11 CBr CH CH CH 12 CCl CH CH CH 13 N CH CCl CH 14 CH CCl CCl CH 15 CH CF CH CH 16 CH CMe CH CH 17 CH N CH CH 18 CH CH CBr CH 19 CH CH CCF₃ CH 20 CH CH CEt CH 21 CH CH CF CH 22 CH CH CMe CH 23 CH CH C—OMe CH 24 CH CH CCl CCl 25 CH CH CH CF 26 CH CH CH C—OMe 27 CH CH CH N 28 Cl CH CH CH 29 CH CH CCN CH 30 CBr CH CCl CH


8. A compound according to any one of claims 1 to 7, or a pharmaceutically acceptable salt, solvate, prodrug, isomer, tautomer, polymorph and/or N-oxide thereof, wherein one, two or three of R^(AA), R^(BB), R^(CC) and R^(DD), (if present) are selected from H, F, Cl, Br, Me, CF₃, cyclopropyl, cyano and OMe, the remainder (if present) are H.
 9. A compound according to any one of claims 1 to 7, or a pharmaceutically acceptable salt, solvate, prodrug, isomer, tautomer, polymorph and/or N-oxide thereof, wherein A¹¹, A¹², A¹³ and A¹⁴ are selected from combinations 1-30: Combination A¹¹ A¹² A¹³ A¹⁴  1 CF CH CH CH  2 CH CBr CH CH  3 CH CCl CH CH  4 CF CCl CH CH  5 CH CH CH CBr  6 CH CH CCl CH  7 CH CF CF CH  8 CH CH CH CH  9 CF CH CF CH 10 CF CH CCl CH 11 CBr CH CH CH 12 CCl CH CH CH 13 N CH CCl CH 14 CH CCl CCl CH 15 CH CF CH CH 16 CH CMe CH CH 17 CH N CH CH 18 CH CH CBr CH 19 CH CH CCF₃ CH 20 CH CH CEt CH 21 CH CH CF CH 22 CH CH CMe CH 23 CH CH C—OMe CH 24 CH CH CCl CCl 25 CH CH CH CF 26 CH CH CH C—OMe 27 CH CH CH N 28 Cl CH CH CH 29 CH CH CCN CH 30 CBr CH CCl CH


10. A compound according to any one of claims 1 to 9, or a pharmaceutically acceptable salt, solvate, prodrug, isomer, tautomer, polymorph and/or N-oxide thereof, wherein R^(C11)=R^(C1), R^(C13)=R^(C3) and R^(C14)=R^(C4).
 11. A compound according to any one of claims 1 to 10, or a pharmaceutically acceptable salt, solvate, prodrug, isomer, tautomer, polymorph and/or N-oxide thereof, wherein R^(C1), A^(C3) and R^(C4) are selected from combinations 1-4: Combination R^(C1) R^(C3) R^(C4) 1 H H H 2 H H F 3 OMe H H 4 OMe H F


12. A compound according to any one of claims 1 to 11, or a pharmaceutically acceptable salt, solvate, prodrug, isomer, tautomer, polymorph and/or N-oxide thereof, wherein R^(C11), R^(C13) and R^(C14) are selected from combinations 1-4: Combination R^(C11) R^(C13) R^(C14) 1 H H H 2 H H F 3 OMe H H 4 OMe H F


13. A compound according to any one of claims 1 to 12, or a pharmaceutically acceptable salt, solvate, prodrug, isomer, tautomer, polymorph and/or N-oxide thereof, wherein Y is (CH₂)₂.
 14. A compound according to any one of claims 1 to 13, or a pharmaceutically acceptable salt, solvate, prodrug, isomer, tautomer, polymorph and/or N-oxide thereof, wherein R¹ and R¹¹ are selected from C(═O)OH, tetrazolyl and oxo-oxadiazolyl.
 15. A compound according to any one of claims 1-14, selected from: 6,6′-(Ethane-1,2-diylbis(5-carbamoyl-4-methoxy-1H-benzo[d]imidazole-1,2-diyl))bis(3-chlorobenzoic acid) (1) 2,2′-(Ethane-1,2-diylbis(5-carbamoyl-1H-benzo[d]imidazole-1,2-diyl))dibenzoic acid (2) 2,2′-(Ethane-1,2-diylbis(5-carbamoyl-1H-benzo[d]imidazole-1,2-diyl))bis(4-bromobenzoic acid) (3) 2,2′-(Ethane-1,2-diylbis(5-carbamoyl-4-methoxy-1H-benzo[d]imidazole-1,2-diyl))dibenzoic acid (4) 2-(2-(1H-Tetrazol-5-yl)phenyl)-1-(2-(2-(2-(2H-tetrazol-5-yl)phenyl)-5-carbamoyl-1H-benzo[d]imidazol-1-yl)ethyl)-1H-benzo[d]imidazole-5-carboxamide (5) 1,1′-(Ethane-1,2-diyl)bis(2-(2-(2H-tetrazol-5-yl)phenyl)-4-methoxy-1H-benzo[d]imidazole-5-carboxamide) (6) 2,2′-(Ethane-1,2-diylbis(5-carbamoyl-4-methoxy-1H-benzo[d]imidazole-1,2-diyl))bis(4-chlorobenzoic acid) (7) 6,6′-(Ethane-1,2-diylbis(5-carbamoyl-4-methoxy-1H-benzo[d]imidazole-1,2-diyl))bis(3,4-dichlorobenzoic acid) (8) 1,1′-(Ethane-1,2-diyl)bis(2-(4-chloro-2-(2H-tetrazol-5-yl)phenyl)-4-methoxy-1H-benzo[d]imidazole-5-carboxamide) (9) 2,2′-(Ethane-1,2-diylbis(5-carbamoyl-4-methoxy-1H-benzo[d]imidazole-1,2-diyl))bis(3-fluorobenzoic acid) (10) 2,2′-(ethane-1,2-diylbis(5-carbamoyl-4-methoxy-1H-benzo[d]imidazole-1,2-diyl))bis(5-chloro-3-fluorobenzoic acid) (11) 2,2′-(Ethane-1,2-diylbis(5-carbamoyl-4-methoxy-1H-benzo[d]imidazole-1,2-diyl))bis(4-chloro-3-fluorobenzoic acid) (12) Dimethyl 6,6′-(ethane-1,2-diylbis(5-carbamoyl-4-methoxy-1H-benzo[d]imidazole-1,2-diyl))bis(3-bromobenzoate) (13) 6,6′-(Ethane-1,2-diylbis(5-carbamoyl-4-methoxy-1H-benzo[d]imidazole-1,2-diyl))bis(3-bromobenzoic acid) (14) 6,6′-(Ethane-1,2-diylbis(5-carbamoyl-4-methoxy-1H-benzo[d]imidazole-1,2-diyl))bis(3,4-difluorobenzoic acid) (15) 2,2′-(Ethane-1,2-diylbis(5-carbamoyl-4-methoxy-1H-benzo[d]imidazole-1,2-diyl))bis(3,5-difluorobenzoic acid) (16) 1,1′-(Ethane-1,2-diyl)bis(2-(4-chloro-2-(5-oxo-4,5-dihydro-1,3,4-oxadiazol-2-yl)phenyl)-4-methoxy-1H-benzo[d]imidazole-5-carboxamide) (17) 2,2′-(Ethane-1,2-diylbis(5-carbamoyl-7-fluoro-4-methoxy-1H-benzo[d]imidazole-1,2-diyl))bis(3,5-difluorobenzoic acid) (18) 2,2′-(Ethane-1,2-diylbis(5-carbamoyl-4-methoxy-1H-benzo[d]imidazole-1,2-diyl))bis(3-iodobenzoic acid) (19.1 and 19.2) 6,6′-(Ethane-1,2-diylbis(5-carbamoyl-7-fluoro-4-methoxy-1H-benzo[d]imidazole-1,2-diyl))bis(3-chlorobenzoic acid) (20) Dimethyl 6,6′-(ethane-1,2-diylbis(5-carbamoyl-4-methoxy-1H-benzo[d]imidazole-1,2-diyl))bis(2-bromobenzoate) (21) 2,2′-(Ethane-1,2-diylbis(5-carbamoyl-4-methoxy-1H-benzo[d]imidazole-1,2-diyl))bis(3-chlorobenzoic acid) (22) 1,1′-(Ethane-1,2-diyl)bis(2-(2-chloro-6-(2H-tetrazol-5-yl)phenyl)-4-methoxy-1H-benzo[d]imidazole-5-carboxamide) (23) 1,1′-(Ethane-1,2-diyl)bis(2-(4-chloro-2-(2H-1,2,3-triazol-4-yl)phenyl)-4-methoxy-1H-benzo[d]imidazole-5-carboxamide) (24) 6,6′-(Ethane-1,2-diylbis(5-carbamoyl-4-methoxy-1H-benzo[d]imidazole-1,2-diyl))bis(3-methylbenzoic acid) (25) 2,2′-(Ethane-1,2-diylbis(5-carbamoyl-4-methoxy-1H-benzo[d]imidazole-1,2-diyl))bis(4-methylbenzoic acid) (26) 6,6′-(Ethane-1,2-diylbis(5-carbamoyl-4-methoxy-1H-benzo[d]imidazole-1,2-diyl))bis(3-fluorobenzoic acid) (27) 2,2′-(Ethane-1,2-diylbis(5-carbamoyl-4-methoxy-1H-benzo[d]imidazole-1,2-diyl))bis(4-fluorobenzoic acid) (28) 2,2′-(Ethane-1,2-diylbis(5-carbamoyl-1H-benzo[d]imidazole-1,2-diyl))bis(3-bromobenzoic acid) (29.1 and 29.2) 3,3′-(Ethane-1,2-diylbis(5-carbamoyl-4-methoxy-1H-benzo[d]imidazole-1,2-diyl))diisonicotinic acid (30) 2,2′-(Ethane-1,2-diylbis(5-carbamoyl-4-methoxy-1H-benzo[d]imidazole-1,2-diyl))bis(5-chloronicotinic acid) (31) 1,1′-(Ethane-1,2-diyl)bis(2-(4-chloro-2-(5-oxo-4,5-dihydro-1,2,4-oxadiazol-3-yl)phenyl)-4-methoxy-1H-benzo[d]imidazole-5-carboxamide) (32) 6,6′-(Ethane-1,2-diylbis(5-carbamoyl-4-methoxy-1H-benzo[d]imidazole-1,2-diyl))bis(3-ethylbenzoic acid) (33) (E)-6,6′-(But-2-ene-1,4-diylbis(5-carbamoyl-4-methoxy-1H-benzo[d]imidazole-1,2-diyl))bis(3-chlorobenzoic acid (34) 6,6′-(Propane-1,3-diylbis(5-carbamoyl-4-methoxy-1H-benzo[d]imidazole-1,2-diyl))bis(3-chlorobenzoic acid) (35) 6,6′-(Ethane-1,2-diylbis(5-carbamoyl-4-methoxy-1H-benzo[d]imidazole-1,2-diyl))bis(2-bromobenzoic acid) (36) 1,1′-(Ethane-1,2-diyl)bis(2-(3,4-dichloro-2-(2H-tetrazol-5-yl)phenyl)-4-methoxy-1H-benzo[d]imidazole-5-carboxamide) (37) 6,6′-(Ethane-1,2-diylbis(5-carbamoyl-4-methoxy-1H-benzo[d]imidazole-1,2-diyl))bis(3-(trifluoromethyl)benzoic acid) (38) 6,6′-(Ethane-1,2-diylbis(5-carbamoyl-1H-benzo[d]imidazole-1,2-diyl))bis(3-chlorobenzoic acid) (39) 1,1′-(Ethane-1,2-diyl)bis(2-(3-chloro-2-(2H-tetrazol-5-yl)phenyl)-4-methoxy-1H-benzo[d]imidazole-5-carboxamide) (40) 6,6′-(Ethane-1,2-diylbis(5-carbamoyl-4-methoxy-1H-benzo[d]imidazole-1,2-diyl))bis(3-methoxybenzoic acid) (41) 1,1′-(Ethane-1,2-diyl)bis(2-(5-chloro-2-(2H-tetrazol-5-yl)phenyl)-4-methoxy-1H-benzo[d]imidazole-5-carboxamide) (42) 1,1′-(Ethane-1,2-diyl)bis(2-(2,4-difluoro-6-(2H-tetrazol-5-yl)phenyl)-4-methoxy-1H-benzo[d]imidazole-5-carboxamide) (43) 1,1′-(Ethane-1,2-diyl)bis(2-(2,4-difluoro-6-(5-oxo-4,5-dihydro-1,3,4-oxadiazol-2-yl)phenyl)-4-methoxy-1H-benzo[d]imidazole-5-carboxamide) (44) 1,1′-(Ethane-1,2-diyl)bis(2-(2-bromo-4-chloro-6-fluorophenyl)-4-methoxy-1H-benzo[d]imidazole-5-carboxamide) (45) 1,1′-(Ethane-1,2-diyl)bis(2-(4-chloro-2-fluoro-6-(2H-tetrazol-5-yl)phenyl)-4-methoxy-1H-benzo[d]imidazole-5-carboxamide) (46) 2-(5-Carbamoyl-1-(2-(5-carbamoyl-2-(4-chloro-2-(2H-tetrazol-5-yl)phenyl)-4-methoxy-1H-benzo[d]imidazol-1-yl)ethyl)-4-methoxy-1H-benzo[d]imidazol-2-yl)-5-chloro-3-fluorobenzoic acid (47) 1,1′-(Ethane-1,2-diyl)bis(2-(2-fluoro-6-(2H-tetrazol-5-yl)phenyl)-4-methoxy-1H-benzo[d]imidazole-5-carboxamide) (48) 1,1′-(Ethane-1,2-diyl)bis(2-(2-fluoro-6-(5-oxo-4,5-dihydro-1,3,4-oxadiazol-2-yl)phenyl)-4-methoxy-1H-benzo[d]imidazole-5-carboxamide) (49) 1,1′-(Ethane-1,2-diyl)bis(2-(4-chloro-2-fluoro-6-(5-oxo-4,5-dihydro-1,3,4-oxadiazol-2-yl)phenyl)-4-methoxy-1H-benzo[d]imidazole-5-carboxamide) (50) 2-(5-Carbamoyl-1-(2-(5-carbamoyl-2-(2-carboxy-4-cyanophenyl)-4-methoxy-1H-benzo[d]imidazol-1-yl)ethyl)-4-methoxy-1H-benzo[d]imidazol-2-yl)-5-cyanobenzoic acid (51) Methyl 2-(5-carbamoyl-1-(2-(5-carbamoyl-2-(4-cyano-2-(methoxycarbonyl)phenyl)-4-methoxy-1H-benzo[d]imidazol-1-yl)ethyl)-4-methoxy-1H-benzo[d]imidazol-2-yl)-5-cyanobenzoate (52) or a pharmaceutically acceptable salt, solvate, prodrug, isomer, tautomer, polymorph and/or N-oxide thereof.
 16. A compound as defined in any one of claims 1 to 15, or a pharmaceutically acceptable salt, solvate, prodrug, isomer, tautomer, polymorph and/or N-oxide thereof, for use in a method of therapy.
 17. A pharmaceutical composition comprising a compound as defined in any one of claims 1 to 15, or a pharmaceutically acceptable salt, solvate, prodrug, isomer, tautomer, polymorph and/or N-oxide thereof, and a pharmaceutically acceptable excipient.
 18. A compound as defined in any one of claims 1 to 15, or a pharmaceutically acceptable salt, solvate, prodrug, isomer, tautomer, polymorph and/or N-oxide thereof, or pharmaceutical composition according to claim 17 for use in the treatment or prevention of a disease ameliorated by the modulation of STING.
 19. A method of treating or preventing a disease ameliorated by the modulation of STING, comprising administering to a subject in need thereof a therapeutically effective amount of a compound as defined in any one of claims 1 to 15, or a pharmaceutically acceptable salt, solvate, prodrug, isomer, tautomer, polymorph and/or N-oxide thereof, or pharmaceutical composition according to claim
 17. 20. Use of a compound as defined in any one of claims 1 to 15, or a pharmaceutically acceptable salt, solvate, prodrug, isomer, tautomer, polymorph and/or N-oxide thereof, in the preparation of a medicament for treating or preventing a disease ameliorated by the modulation of STING. 